ES2674600T3 - Human antibodies against PCSK9 for use in methods of treating particular subject groups - Google Patents

Abstract

Pharmaceutical composition comprising an antibody or an antigen-binding fragment thereof that specifically binds to proprotein convertase subtilisin / human type 9 kexin (PCSK9h) together with a pharmaceutically acceptable carrier or excipient for use in the treatment of a disease or condition. , in which the expression or activity of PCSK9 causes an impact, in a human patient, in which the antibody or antigen binding fragment thereof comprises the three heavy chain CDRs set forth in SEQ ID NO: 76, 78, and 80 and the three light chain CDRs set forth in SEQ ID NO: 84, 86, and 88, wherein the pharmaceutical composition is comprised in a unit dosage form, wherein the unit dosage form comprises 75 mg, 150 mg or 300 mg of the antibody or antigen binding fragment thereof, in which the pharmaceutical composition is administered every two to four weeks, and in which the disease or condition is selected. from the group consisting of high levels of total cholesterol, high levels of non-high-density lipoprotein (HDL) cholesterol, high levels of low-density lipoprotein (C-LDL), high levels of apolipoprotein B100 (ApoB100), hyperlipidemia, dyslipidemia, atherosclerosis, cardiovascular diseases, such as coronary heart disease (CHD) and hypercholesterolemia, such as primary hypercholesterolemia, familial hypercholesterolemia, unfamiliar hypercholesterolemia, heterozygous familial hypercholesterolemia (heFH), hypercholesterolemia that is not controlled by statins, and hypercholesterolemia that is resistant to statins

Description

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DESCRIPTION

Human antibodies against PCSK9 for use in methods of treating particular subject groups

The present invention relates to pharmaceutical compositions comprising antibodies specific to PCSK9 or antigen binding fragments thereof for use in the treatment of diseases or conditions in which the expression or activity of proprotein convertase subtilisin / kexin type 9 ( PCSK9) causes an impact. Antibodies specific to PCSK9 or antigen binding fragments thereof (preferably in combination with HMG-CoA reductase inhibitors) are also disclosed for use in the treatment of diseases or conditions in which the expression or activity of PCSK9 causes an impact. .

Additionally, manufacturing articles are disclosed comprising packaging material, PCSK9-specific antibodies or antigen binding fragments thereof, and a label or leaflet indicating which groups of patients can be treated with said antibodies or fragments, which groups of patients they should not be treated with such antibodies or fragments, and what dosage schedule should be used.

Additionally, methods for testing the efficacy of specific antibodies for PCSK9 or antigen binding fragments thereof for the treatment of certain diseases or conditions and for the treatment of specific subgroups of patients are disclosed.

BACKGROUND OF THE INVENTION

Proprotein convertase subtilisin / kexin type 9 (PCSK9) is a proprotein convertase that belongs to the subfamily of proteinase K of the family of secretory subtylases. The encoded protein is synthesized as a soluble zymogen that undergoes autocatalytic intramolecular processing in the endoplasmic reticulum. Evidence suggests that PCSK9 increases LDL cholesterol in plasma by promoting the degradation of the LDL receptor, which mediates the endocytosis of LDL in the liver, the main route of elimination of LDL from the circulation. The structure of the PCSK9 protein shows that it has a signal sequence, followed by a dominance, a catalytic domain that contains a conserved triad of residues (D186, H226 and S386) and a C-end domain. It is synthesized as a 74 kDa precursor. soluble that undergoes autocatalytic excision in the ER, generating a dominance of 14 kDa and catalytic fragment of 60 kDa. It has been shown that autocatalytic activity is required for secretion. After cleavage, the dominance remains strongly associated with the catalytic domain.

Antibodies against PCSK9 are described in, for example, WO 2008/057457, WO 2008/057458, WO 2008/057459, WO 2008/063382, WO 2008/125623 and US 2008/0008697. Anti-PCSK9 antibodies that are particularly very suitable for the practice of the present invention are disclosed in US 2010/0166768 A1.

Amgen: "Ascending Multiple Dose Study to Evaluate the Safety, Tolerability, Pharmacokinetics and Pharmacodynamics of AMG 145 in Subjects With Hyperlipidemia on Stable Doses of a Statin", May 27, 2010 and Pfizer: "Safety and Tolerability of Multiple Doses Of PF- 04950615 (RN316) In Subjects With Hypercholesterolemia, "November 3, 2010, both retrieved from the clinicaltrials.gov database, studied the effect of AMG145 and RN316, respectively.

WO 2010/029513 A2 discloses antagonists of PCSK9.

WO 2009/026558 A1 discloses antigen-binding proteins for PCSK9.

WO 2009/055783 A2 discloses anti-PCSK9 antibodies or antigen-binding fragments thereof and methods of treating lipid and cholesterol disorders.

WO 2012/054438 A1 discloses an anti-PCSK9 antibody and methods of treating medical disorders, such as elevated LDL, using such an antibody.

WO 2012/064792 A2 discloses protein complexes for antigen binding and methods of use.

Soutar A. K. (Current Opinion in Lipidology, vol. 22, No. 3, June 2011, p. 192-196) reveals the unexpected function for PCSK9 in lipid metabolism.

Joyce C.Y. Chan et al. (Proceedings in the national academy of sciences, vol. 106, No. 24, June 16, 2009) disclose a neutralizing antibody against PCSK9 that lowers serum cholesterol in mice and nonhuman primates.

UNDERLYING TECHNICAL PROBLEMS TO THE PRESENT INVENTION

Statins are among the most widely used drugs in the world. Although statins generally have an excellent safety profile, it is desired to further optimize the safety profile by reducing the already low rate of unwanted side effects (such as myopathies).

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Despite the widespread availability of lipid lowering agents such as statins, approximately 30% of all adult patients treated for hypercholesterolemia in the United States between 1999 and 2006 failed to achieve their recommended LDL-C targets. Reasons for this include poor therapeutic compliance with therapy, drug resistance / intolerance and the positive relationship between adverse event rates and dose increase. In addition, since the most effective lipid lowering agents can only reduce LDL-C levels up to 55%, the rates of obtaining targets in patients requiring substantial reductions in LDL-C, such as those with familial hypercholesterolemia, are often significantly more casualties than might be expected. Therefore, more effective lipid lowering agents and treatment guidelines are required to improve the rates of obtaining targets in these patients.

Surprisingly enough, the inventors of the present invention found that the administration of anti-PCSK9 antibodies or fragments thereof increases the LDL cholesterol lowering activity of statins, when administered in particular dosage schedules and / or to particular groups of patients.

Thus, the co-administration of anti-PCSK9 antibodies or fragments thereof enhances the effectiveness of a statin therapy and allows a reduction in the statin dosage, thus reducing unwanted side effects.

In addition, the inventors of the present invention found that particular dosage regimens of anti-PCSK9 antibodies and / or statins are more suitable for lowering LDL cholesterol levels than others. The inventors also found that some subgroups of patients benefit more than others from treatment with anti-PCSK9 antibodies or fragments thereof and / or statins. The inventors also found that treatment with anti-PCSK9 antibodies or fragments thereof and / or statins is contraindicated for some subgroups of patients.

The above overview does not necessarily describe all the problems solved by the present invention. SUMMARY OF THE INVENTION

The scope of the present invention is defined in the attached set of claims.

In a first aspect, the present disclosure refers to a method of treatment of a disease or condition in which the expression or activity of PCSK9 causes an impact, comprising:

administering a therapeutic amount of an antibody or antigen-binding fragment thereof that specifically binds PCSK9h (proprotein convertase subtilisin / human type 9 kexin) to a subject in need thereof, in which the antibody or fragment binding to the antigen thereof is administered in a dose amount ranging from 5 mg to 500 mg, and

administering a therapeutic amount of an HMG-CoA reductase inhibitor to said subject, wherein the HMG-CoA reductase inhibitor is administered in a dose amount ranging from 0.05 mg to 100 mg.

In a second aspect, the present disclosure relates to an antibody or an antigen-binding fragment thereof that specifically binds PCSK9h (proprotein convertase subtilisin / human type 9 kexin) for use in the treatment of a disease or condition in that the expression or activity of PCSK9 causes an impact,

wherein the antibody or antigen binding fragment thereof is for administration in a dose amount ranging from 5 mg to 500 mg,

wherein the antibody or antigen binding fragment thereof is also for administration in combination with an HMG-CoA reductase inhibitor at a dose amount ranging from 0.05 mg to 100 mg.

In a third aspect, the present disclosure refers to an article of manufacture comprising:

(a) a packaging material; (b) an antibody or an antigen binding fragment thereof that specifically binds to PCSK9h; and (c) a label or leaflet contained within the packaging material that indicates that patients receiving treatment with said antibody or antigen binding fragment can be treated for a disease or condition selected from the group consisting of hypercholesterolemia, hyperlipidemia, dyslipidemia, atherosclerosis and cardiovascular diseases.

In a fourth aspect, the present disclosure refers to an article of manufacture comprising:

(a) a packaging material; (b) an antibody or an antigen binding fragment thereof that specifically binds to PCSK9h; and (c) a label or leaflet contained within the packaging material that indicates the treatment of patients with said antibody or antigen binding fragment thereof together with the application of a statin.

In a fifth aspect, the present disclosure refers to an article of manufacture comprising (a) a packaging material; (b) an antibody or an antigen binding fragment thereof that specifically binds to

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PCSK9h; and (c) a label or leaflet indicating that the treatment of patients with said antibody or antigen-binding fragment thereof together with a statin is contraindicated for patients belonging to one or more of the following groups: (i) smokers; (ii) persons 70 years of age or older; (iii) people suffering from hypertension; (iv) women who are pregnant; (v) women who are trying to get pregnant; (vi) women who are breastfeeding; (vii) people who have or have ever had a disease that affects the liver; (viii) people who had some unexplained abnormal blood test for liver function; (ix) people who drink excessive amounts of alcohol; (x) people who have kidney problems; (xi) people suffering from hypothyroidism; (xii) people suffering from muscular disorders; (xiii) people who have had previous muscle problems during treatment with lipid lowering medicine; (xiv) people who have serious problems with their breathing; (xv) people taking one or more of the following medicines: medicines that alter the way the immune systems work (for example, cyclosporine or antihistamines), antibiotics or antifungal medicines (for example, erythromycin, clarithromycin, ketoconazole, itraconazole , rifampicin, fusidic acid), medicines that regulate lipid levels (for example, gemfibrozil, colestipol), calcium channel blockers (for example, verapamil, diltiazem), medicines that regulate heart rhythm (digoxin, amiodarone), protease inhibitors used in the treatment of HIV (for example, nelfinavir), warfarin, oral contraceptives, antacids or St. John's wort; or (xvi) people who drink more than 0.1 l of grapefruit juice a day; (xvii) people who have a body mass index (BMI) greater than 40; (xviii) people who have a body mass index (BMI) of less than 18; (xix) people suffering from type 1 diabetes or type 2 diabetes; (xx) people positive for hepatitis B or hepatitis C; or (xxi) people who have a known sensitivity to therapeutic drugs of monoclonal antibodies.

In a sixth aspect, the present disclosure relates to a method of testing the efficacy of an antibody or an antigen binding fragment thereof that specifically binds to PCSK9h for the treatment of a disease or condition selected from the group consisting of hypercholesterolemia, hyperlipidemia, dyslipidemia, atherosclerosis and cardiovascular diseases, said method comprising:

treating a selected patient population with said antibody or antigen binding fragment thereof, in which each patient in said population has a C-LDL cholesterol level greater than 100 mg / dl; Y

determining the efficacy of said antibody or antigen binding fragment thereof by determining the level of LDL-C in the patient population before and after administration of said antibody or antigen binding fragment thereof, in which a reduction in the LDL-C level of at least 25% with respect to a predose level in at least 75% of the patient population indicates that said antibody or antigen-binding fragment thereof is effective for the treatment of said disease or condition in said patient population.

In a seventh aspect, the present disclosure relates to a method of testing the efficacy of an antibody or an antigen binding fragment thereof that specifically binds to PCSK9h for the treatment of a disease or condition selected from the group consisting of hypercholesterolemia, hyperlipidemia, dyslipidemia, atherosclerosis and cardiovascular diseases, said method comprising:

determining the efficacy of an antibody or antigen binding fragment thereof that has been used for the treatment of a selected patient population with said antibody or antigen binding fragment thereof, in which each patient in said population has a level of C-LDL cholesterol greater than 100 mg / dl determining the level of C-LDL in the patient population before and after the administration of said antibody or antigen-binding fragment thereof, in which a reduction in the level of LDL-C of at least 25% with respect to a predose level in at least 75% of the patient population indicates that said antibody or antigen-binding fragment thereof is effective for the treatment of said disease or condition in said patient population

In an eighth aspect, the present disclosure relates to a package comprising an antibody or antigen binding fragment thereof that specifically binds to PCSK9h (see "Preferred antibodies" section) and a label, said label comprising a printed statement which informs the patient that the treatment of the antibody together with a statin is indicated in one or more of the indications selected from the group consisting of hypercholesterolemia, hyperlipidemia, dyslipidemia, atherosclerosis and cardiovascular diseases.

In a ninth aspect, the present disclosure relates to a package comprising an antibody or antigen binding fragment thereof that specifically binds to PCSK9h (see section "Preferred Antibodies") and a label, said label comprising a printed statement that informs the patient that the treatment of the antibody together with a statin is contraindicated for patients belonging to one or more of the following groups: (i) smokers; (ii) persons 70 years of age or older; (iii) people suffering from hypertension; (iv) women who are pregnant; (v) women who are trying to get pregnant; (vi) women who are breastfeeding; (vii) people who have or have ever had a disease that affects the liver; (viii) people who had some unexplained abnormal blood test for liver function; (ix) people who drink excessive amounts of alcohol; (x) people who have kidney problems; (xi) people suffering from hypothyroidism; (xii) people suffering from muscular disorders; (xiii) people who have had previous muscle problems during

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treatment with lipid lowering medicine; (xiv) people who have serious problems with their breathing; (xv) people taking one or more of the following medicines: medicines that alter the way the immune systems work (for example, cyclosporine or antihistamines), antibiotics or antifungal medicines (for example, erythromycin, clarithromycin, ketoconazole, itraconazole , rifampicin, fusidic acid), medicines that regulate lipid levels (for example, gemfibrozil, colestipol), calcium channel blockers (for example, verapamil, diltiazem), medicines that regulate heart rhythm (digoxin, amiodarone), protease inhibitors used in the treatment of HIV (for example, nelfinavir), warfarin, oral contraceptives, antacids or St. John's wort; or (xvi) people who drink more than 0.1 l of grapefruit juice a day; (xvii) people who have a body mass index (BMI) greater than 40; (xviii) people who have a body mass index (BMI) of less than 18; (xix) people suffering from type 1 diabetes or type 2 diabetes; (xx) people positive for hepatitis B or hepatitis C; or (xxi) people who have a known sensitivity to therapeutic drugs of monoclonal antibodies.

In a tenth aspect, the present disclosure relates to a method of regulating the level of LDL in the blood comprising:

administering a therapeutic amount of an antibody or antigen-binding fragment thereof that specifically binds PCSK9h (proprotein convertase subtilisin / human type 9 kexin) to a subject in need thereof, in which the antibody or fragment binding to the antigen thereof is administered in a dose amount ranging from 5 mg to 500 mg, and

administering a therapeutic amount of an HMG-CoA reductase inhibitor to said subject, wherein the HMG-CoA reductase inhibitor is administered in a dose amount ranging from 0.05 mg to 100 mg.

In a eleventh aspect, the present disclosure relates to a method of preventing the effects of a (persistently) high level of LDL in the blood comprising:

administering a therapeutic amount of an antibody or antigen-binding fragment thereof that specifically binds PCSK9h (proprotein convertase subtilisin / human type 9 kexin) to a subject in need thereof, in which the antibody or fragment binding to the antigen thereof is administered in a dose amount ranging from 5 mg to 500 mg, and

administering a therapeutic amount of an HMG-CoA reductase inhibitor to said subject, wherein the HMG-CoA reductase inhibitor is administered in a dose amount ranging from 0.05 mg to 100 mg.

In a twelfth aspect, the present disclosure relates to a method of determining whether a pharmaceutical compound is usable to correct, improve, inhibit or prevent a disease or condition in which the activity or expression of PCSK9 has an impact comprising:

(a) administering to a subject a compound that specifically binds PCSK9, preferably an antibody or antigen binding fragment thereof that specifically binds to PCSK9, and

(b) determine which fraction of PCSK9 in the blood is bound to the compound of (a).

In a thirteenth aspect, the present disclosure relates to a method of treatment of a disease or condition in which the expression or activity of PCSK9 causes an impact comprising

administering a therapeutic amount of an antibody or an antigen-binding fragment thereof that specifically binds PCSK9h (proprotein convertase subtilisin / human type 9 kexin) to a subject in need thereof,

in which the subject in need thereof is classified into one or more of the following groups of subjects: (i) subjects who have a serum LDL-C-cholesterol level of at least 100 mg / dl; (ii) subjects who have a serum HDL-C level below 40 mg / dl; (iii) subjects who have a serum cholesterol level of at least 200 mg / dl; (iv) subjects who have a serum triacylglycerols level of at least 150 mg / dl, wherein said triacylglycerols level is determined after fasting for at least 8 hours; (v) subjects who are at least 35 years old; (vi) subjects younger than 75 years; (vii) subjects who have a BMI of 25 or more; (viii) male subjects; (ix) female subjects; (x) subjects in which the administration of said antibody or antigen binding fragment thereof leads to a reduction in the level of serum LDL-C of at least 30 mg / dl with respect to the predose level; or (xi) subjects in which the administration of said antibody or antigen binding fragment thereof leads to a reduction in the level of serum LDL-C of at least 20% with respect to the predose level.

In a fourteenth aspect, the present disclosure refers to a method of treatment of a disease or condition in which the expression or activity of PCSK9 causes an impact comprising

administering a therapeutic amount of an antibody or an antigen-binding fragment thereof that specifically binds PCSK9h (proprotein convertase subtilisin / human type 9 kexin) to a subject in need thereof,

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in which the subject in need thereof is not classified into one or more of the following groups of subjects: (i) smokers; (ii) persons 70 years of age or older; (iii) people suffering from hypertension; (iv) women who are pregnant; (v) women who are trying to get pregnant; (vi) women who are breastfeeding; (vii) people who have or have ever had a disease that affects the liver; (viii) people who had some unexplained abnormal blood test for liver function; (ix) people who drink excessive amounts of alcohol; (x) people who have kidney problems; (xi) people suffering from hypothyroidism; (xii) people suffering from muscular disorders; (xiii) people who have had previous muscle problems during treatment with lipid lowering medicine; (xiv) people who have serious problems with their breathing; (xv) people taking one or more of the following medicines: medicines that alter the way the immune systems work (for example, cyclosporine or antihistamines), antibiotics or antifungal medicines (for example, erythromycin, clarithromycin, ketoconazole, itraconazole , rifampicin, fusidic acid), medicines that regulate lipid levels (for example, gemfibrozil, colestipol), calcium channel blockers (for example, verapamil, diltiazem), medicines that regulate heart rhythm (digoxin, amiodarone), protease inhibitors used in the treatment of HIV (for example, nelfinavir), warfarin, oral contraceptives, antacids or St. John's wort; or (xvi) people who drink more than 0.1 l of grapefruit juice a day; (xvii) people who have a body mass index (BMI) greater than 40; (xviii) people who have a body mass index (BMI) of less than 18; (xix) people suffering from type 1 diabetes or type 2 diabetes; (xx) people positive for hepatitis B or hepatitis C; or (xxi) people who have a known sensitivity to therapeutic drugs of monoclonal antibodies.

In a fifteenth aspect, the present disclosure relates to an antibody or an antigen-binding fragment thereof that specifically binds PCSK9h (proprotein convertase subtilisin / human type 9 kexin), in which the antibody is characterized by one or more more of the following characteristics after administration to a subject, preferably a human or non-human mammal:

1. Reduction of low-density lipoprotein (C-LDL) levels from at least about -25% to about -40% compared to a predose level with a sustained reduction for at least a period of 14 days, in wherein the sustained reduction is preferably at least -25% and more preferably at least -30% with respect to a predose level, particularly if administered in a dose of about 40 to about 60 mg, preferably about 45 to about 55 mg and more preferably about 50 mg in a biweekly administration schedule (every two weeks, E2W),

2. Reduction of low density lipoprotein (C-LDL) from at least about -50% to about -65% with respect to a predose level with a sustained reduction for at least a period of 14 days, in which the Sustained reduction is preferably at least -40% and more preferably at least -45% with respect to a predose level, particularly if administered in a dose of about 100 mg E2W,

3. Reduction of low density lipoprotein (C-LDL) from at least about -60% to at least about -75% [eg, at least about -60%, at least about -65%, at less about -70 or at least about -75%] with respect to a predose level with a sustained reduction for at least a period of 14 days, in which the sustained reduction is preferably at least -55% and more preferably at least -60% with respect to a predose level, particularly when administered in a dose of approximately 150 mg E2W,

4. Reduction of low density lipoprotein (C-LDL) of at least about 40% at

approximately 75% with respect to a predose level with a sustained reduction for at least a period of 28 days, in which the sustained reduction is preferably at least -35% and more preferably at least -40% with respect to a predose level, particularly when administered in a dose of approximately 200 mg E4W,

5. Reduction of low density lipoprotein (C-LDL) of at least about -50% at

approximately -75% with respect to a predose level with a sustained reduction for at least a period of 28 days, in which the sustained reduction is preferably at least -40% and more preferably at least -45% with respect to a predose level, particularly when administered in a dose of approximately 300 mg E4W,

6. Increased serum HDL cholesterol levels of at least 2%, at least 2.5%, at least

3%, at least 3.5%, at least 4%, at least 4.5%, at least 5% or at least 5.5% with respect

at a predose level, particularly when administered in a dose of approximately 150 mg E2W,

7. reduction of total serum cholesterol from at least about 25% to about 35% with respect to a predose level with a sustained reduction for at least a 24-day period,

8. reduction of total serum cholesterol from at least about 65% to about 80% with respect to a predose level with a sustained reduction for at least a 24-day period,

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9. reduction of serum triglyceride levels from at least about 25% to about

40% with respect to a predose level,

10. Little or no measurable effect on liver function, as determined by ALT measurements and

AST,

11. little or no measurable effect on troponin levels,

12. increase in one or more of: total cholesterol levels, ApoB levels, C-non HDL levels, Apo- ratio

B / ApoA-1,

In a sixteenth aspect, the present disclosure relates to an antibody or an antigen-binding fragment thereof that specifically binds PCSK9h (proprotein convertase subtilisin / human type 9 kexin) for use in the treatment of a disease or condition wherein the expression or activity of PCSK9 causes an impact, in which the antibody or antigen binding fragment thereof is for administration at a dose of about 50 to 500 mg.

In a seventeenth aspect, the present disclosure relates to an antibody or an antigen binding fragment thereof that specifically binds PCSK9h (proprotein convertase subtilisin / human type 9 kexin).

for use in the treatment of a disease or condition in which the expression or activity of PCSK9 causes an impact.

wherein the antibody or antigen binding fragment thereof is for administration to a subject that is in at least one of the following groups of subjects: (i) subjects having a serum LDL cholesterol level of at least 100mg / dl (i.e. at least 2.6 mmol / l) or at least 115mg / dl (i.e. at least 3.0 mmol / l); (ii) subjects who have a serum C-HDL level below 40 mg / dl; (iii) subjects who have a serum cholesterol level of at least 200 mg / dl; (iv) subjects who have a serum triacylglycerols level of at least 150 mg / dl, wherein said triacylglycerols level is determined after fasting for at least 8 hours; (v) subjects who are at least 18, 24 or 35 years old; (vi) subjects who are 75 years old or younger; (vii) subjects who have a BMI of 25 or more or 30 or more; (viii) male subjects; (ix) female subjects; (x) subjects in which the administration of said antibody or antigen binding fragment thereof leads to a reduction in the level of serum LDL-C of at least 30 mg / dl, at least 40 mg / dl, at least 45 mg / dl or at least 50 mg / dl with respect to the predose level (especially after 12 weeks of treatment); or (xi) subjects in which the administration of said antibody or antigen binding fragment thereof leads to a reduction in the level of serum LDL-C of at least 20%, at least 25%, at least 30 %, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65% or at least 70% with respect at the predose level especially after 12 weeks of treatment).

In an eighteenth aspect, the present disclosure relates to an antibody or antigen binding fragment thereof that specifically binds PCSK9h (proprotein convertase subtilisin / human type 9 kexin) for use in the treatment of a disease or condition in which the expression or activity of PCSK9 causes an impact,

wherein the antibody or antigen binding fragment thereof is for administration to a subject that is not classified into one or more of the following groups of subjects: (i) smokers; (ii) persons 70 years of age or older; (iii) people suffering from hypertension; (iv) women who are pregnant; (v) women who are trying to get pregnant; (vi) women who are breastfeeding; (vii) people who have or have ever had a disease that affects the liver; (viii) people who had some unexplained abnormal blood test for liver function; (ix) people who drink excessive amounts of alcohol; (x) people who have kidney problems; (xi) people suffering from hypothyroidism; (xii) people suffering from muscular disorders; (xiii) people who have had previous muscle problems during treatment with lipid lowering medicine; (xiv) people who have serious problems with their breathing; (xv) people taking one or more of the following medicines: medicines that alter the way the immune systems work (for example, cyclosporine or antihistamines), antibiotics or antifungal medicines (for example, erythromycin, clarithromycin, ketoconazole, itraconazole , rifampicin, fusidic acid), medicines that regulate lipid levels (for example, gemfibrozil, colestipol), calcium channel blockers (for example, verapamil, diltiazem), medicines that regulate heart rhythm (digoxin, amiodarone), protease inhibitors used in the treatment of HIV (for example, nelfinavir), warfarin, oral contraceptives, antacids or St. John's wort; or (xvi) people who drink more than 0.1 l of grapefruit juice a day; (xvii) people who have a body mass index (BMI) greater than 40; (xviii) people who have a body mass index (BMI) of less than 18; (xix) people suffering from type 1 diabetes or type 2 diabetes; (xx) people positive for hepatitis B or hepatitis C; or (xxi) people who have a known sensitivity to therapeutic drugs of monoclonal antibodies.

In the eleventh aspect, the present disclosure relates to a pharmaceutical composition comprising the antibody or antigen-binding fragment thereof disclosed herein together with a pharmaceutically acceptable carrier or excipient.

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In a twentieth aspect, the present disclosure relates to a solution for injection as described herein comprising the antibody or antigen binding fragment thereof disclosed herein, and preferably comprising about 40 mg to about 200 mg or about 50 to about 200 mg, for example, about 40 mg, about 50 mg, about 75 mg, about 100 mg, about 150 mg or about 200 mg of the antibody or antigen binding fragment thereof per 1 ml volume .

In a twenty-first aspect, the present disclosure relates to a dry formulation as described herein comprising the antibody or antigen binding fragment thereof disclosed herein, and preferably comprising about 40 mg to about 500 mg, 50 to about 500 mg, about 50 to about 400, about 50 to about 300 for example, about 40 mg, about 50 mg, about 75 mg, about 100 mg, about 150 mg, about 200 mg, about 250 mg, approximately 300 mg, approximately 350 mg, approximately 400 mg, approximately 450 mg or approximately 500 mg and more preferably approximately 50 mg, approximately 100 mg, approximately 150 mg, approximately 200 mg, approximately 250 mg, approximately 300 mg and even more preferably approximately 150 mg, approximately 200 mg or approximately 300 mg of the antibody or antigen binding fragment thereof by dose.

In a twenty-second aspect, the present disclosure relates to an antibody or antigen-binding fragment thereof comprised in one of the pharmaceutical compositions according to the tenth ninth aspect.

In a twenty-third aspect, the present disclosure relates to a unit dosage form comprising the antibody, antigen-binding fragment thereof or pharmaceutical composition disclosed herein.

In a twenty-fourth aspect, the present disclosure relates to a manufacturing article comprising the pharmaceutical composition disclosed herein, the liquid formulation disclosed herein, or the dry formulation disclosed herein, the antibody or fragment of antigen binding thereof disclosed herein or one or more unit dosage forms disclosed herein and a container or container.

In a twenty-fifth aspect, the present disclosure refers to an article of manufacture comprising: (a) a packaging material; (b) an antibody or an antigen binding fragment thereof that specifically binds to PCSK9h; and (c) a label or leaflet contained within the packaging material that indicates that patients receiving treatment with said antibody or antigen binding fragment can be treated for a disease or condition selected from the group consisting of hypercholesterolemia, hyperlipidemia, dyslipidemia, Atherosclerosis and cardiovascular diseases and additionally indicating that subjects that are in one or more groups of subjects can be treated as cited in the thirteenth aspect.

In a twenty-sixth aspect, the present disclosure refers to an article of manufacture comprising: (a) a packaging material; (b) an antibody or an antigen binding fragment thereof that specifically binds to PCSK9h; and (c) a label or leaflet contained within the packaging material that indicates that patients receiving treatment with said antibody or antigen binding fragment can be treated for a disease or condition selected from the group consisting of hypercholesterolemia, hyperlipidemia, dyslipidemia, Atherosclerosis and cardiovascular diseases and additionally indicating that the treatment of patients with said antibody or antigen binding fragment thereof is contraindicated for patients belonging to one or more groups of subjects as cited in the fourteenth aspect.

In a twenty-seventh aspect, the present disclosure refers to a pharmaceutical composition or antibody or antigen-binding fragment thereof disclosed herein, such as according to the fifteenth or ninth aspect disclosed herein, for use. in the treatment of a disease or condition in which the expression or activity of PCSK9 causes an impact.

In a twenty-eighth aspect, the present disclosure relates to a method of preparing a pharmaceutical composition disclosed herein, for example, according to the nineteenth aspect, comprising mixing the antibody or antigen-binding fragment thereof and optionally the HMG-CoA reductase inhibitor with one or more pharmaceutical excipients or vehicles.

In a twenty-ninth aspect, the present disclosure relates to a method of preparing a unit dosage form comprising administering an amount of the pharmaceutical composition, of the antibody or antigen binding fragment thereof, of the liquid formulation or of the Dry formulation disclosed herein comprising one or more doses of the antibody or antigen fragment thereof and optionally HMG-CoA reductase inhibitor and adapting them as physically discrete units suitable as unit dosages for human and / or animal administration.

In a thirtieth aspect, the present disclosure relates to a method of preparing or assembling an article of manufacture disclosed herein comprising packaging the pharmaceutical composition, of the

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antibody according to, of the liquid formulation, of the dry formulation according to or of or more of the unit dosage forms disclosed herein in a container, optionally together with one or more of the following: a label, instructions for use, An administration device.

In a thirty-first aspect, the present disclosure relates to a method of testing the efficacy of an antibody or an antigen binding fragment thereof that specifically binds to PCSK9h for the treatment of a disease or condition selected from the group consisting of in hypercholesterolemia, hyperlipidemia, dyslipidemia, atherosclerosis and cardiovascular diseases, said method comprising:

treating a selected patient population with said antibody or antigen binding fragment thereof, in which each patient in said population has a C-LDL cholesterol level greater than 100 mg / dl; Y

determining the efficacy of said antibody or antigen binding fragment thereof by determining the level of LDL-C in the patient population before and after administration of said antibody or antigen binding fragment thereof, in which a reduction in the LDL-C level of at least 25% with respect to a predose level in at least 75% of the patient population indicates that said antibody or antigen-binding fragment thereof is effective for the treatment of said disease or condition in said patient population;

in which each patient is classified into one or more groups of subjects as cited in the thirteenth aspect.

In a thirty-second aspect, the present disclosure relates to a method of testing the efficacy of an antibody or an antigen binding fragment thereof that specifically binds to PCSK9h for the treatment of a disease or condition selected from the group consisting of in hypercholesterolemia, hyperlipidemia, dyslipidemia, atherosclerosis and cardiovascular diseases, said method comprising:

determining the efficacy of an antibody or antigen binding fragment thereof that has been used for the treatment of a patient population selected from said antibody or antigen binding fragment thereof, in which each patient in said population has a level of C-LDL cholesterol greater than 100 mg / dl determining the level of C-LDL in the patient population before and after the administration of said antibody or antigen-binding fragment thereof, in which a reduction in the level of LDL-C of at least 25% with respect to a predose level in at least 75% of the patient population indicates that said antibody or antigen-binding fragment thereof is effective for the treatment of said disease or condition in said patient population;

in which each patient is classified into one or more groups of subjects as cited in the thirteenth aspect.

In a thirty-third aspect, the present disclosure relates to a method of testing the efficacy of a compound in reducing cholesterol levels in a subject, comprising the steps: (a) providing a rodent; (b) administering an antibody or an antigen binding fragment thereof that specifically binds PCSK9 to the rodent; (c) administering a test compound to said rodent; (d) determine the effect of the test compound on the rodent, in which a reduction in the level of cholesterol in the rodent compared to the cholesterol level of a control animal indicates that the test compound is effective in reducing the levels of cholesterol in a subject, in which the control animal is of the same species as said rodent, and in which the control animal has not been exposed to the test compound.

In a thirty-fourth aspect, the present disclosure relates to a method of enhancing the activity of LDL-C reduction in a subject receiving statin therapy, the method comprising administering to the subject an antibody, or antigen-binding fragment thereof, which specifically binds to proprotein convertase subtilisin / human type 9 kexin (PCSK9h), in which the antibody or antigen binding fragment thereof is administered in a dose amount within the range of about 5 mg to about 500 mg, thereby enhancing the C-LDL reducing activity of statin therapy in the subject.

In a thirty-fifth aspect, the present disclosure relates to a kit for treating high levels of low density lipoprotein (C-LDL) cholesterol in a subject, the kit comprising (a) unit dosage pharmaceutical form comprising an antibody, or antigen binding fragment thereof, which specifically binds to PCSK9h; and pharmaceutically acceptable carrier, wherein the antibody or antigen-binding fragment is present in a dose amount within the range of about 5 mg to about 500 mg; and (b) a label or leaflet with instructions for its use.

In a thirty-sixth aspect, the present disclosure refers to a method of treating a subject suffering from a disease or disorder characterized by elevated levels of low density lipoprotein cholesterol (LDL-C), the method comprising:

(a) select a subject with a blood LDL-C level greater than 100 mg / dl; Y

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(b) administering to said subject a composition comprising an antibody or antigen binding fragment thereof that specifically binds to proprotein convertase subtilisin / human type 9 kexin (PCSK9h); thus reducing cholesterol levels in the subject in need thereof.

In a twenty-sixth aspect, the present disclosure refers to a method of reducing cholesterol levels in a subject in need thereof, comprising:

(a) select a subject with a low density lipoprotein cholesterol (C-LDL) level in blood greater than 100 mg / dl; Y

(b) administering to said subject a composition comprising an antibody or antigen binding fragment thereof that specifically binds to proprotein convertase subtilisin / human type 9 kexin (PCSK9h); thus reducing cholesterol levels in the subject in need thereof.

Other embodiments will be apparent from a review of the following detailed description.

BRIEF DESCRIPTION OF THE DRAWINGS

Fig. 1 shows the percentage reduction in LDL (C-LDL) cholesterol levels with respect to the initial level for three groups of patients after treatment with the anti-PCSK9 316P antibody. These patient groups are: (1) patients with familial hypercholesterolemia (HeFH); (2) patients with other forms of primary hypercholesterolemia (not FH) with diet and stable atorvastatin therapy; and (3) patients with other forms of primary hypercholesterolemia (not Fh) with diet alone. A dose of 50 mg of the anti-PCSK9 antibody was administered subcutaneously on days 1, 29 and 43. The results of the groups of patients who received the antibody (50-mg-FH-no; 50-mg-FH- Yes; 50-mg-combined) are shown in solid lines, while the results of patients who received a placebo (PBO-FH-no; PBO-FH-Yes; PBO- combined) are shown in dashed lines.

Fig. 2 shows the percentage reduction in LDL (C-LDL) cholesterol levels with respect to the initial level for three groups of patients after treatment with the anti-PCSK9 316P antibody. These patient groups are: (1) patients with familial hypercholesterolemia (HeFH); (2) patients with other forms of primary hypercholesterolemia (not FH) with diet and stable atorvastatin therapy; and (3) patients with other forms of primary hypercholesterolemia (not FH) with diet alone. A dose of 100 mg of the

anti-PCSK9 antibody subcutaneously on days 1, 29 and 43. Results of patient groups

who received the antibody (100-mg-FH-no; 100-mg-FH-Yes; 100-mg-combined) are shown in solid lines, while the results of patients who received a placebo (PBO-FH-no ; PBO-FH-Yes; PBO-combined) are shown in dashed lines.

Fig. 3 shows the percentage reduction in LDL (C-LDL) cholesterol levels with respect to the initial level for three groups of patients after treatment with the anti-PCSK9 316P antibody. These patient groups are: (1) patients with familial hypercholesterolemia (HeFH); (2) patients with other forms of primary hypercholesterolemia (not FH) with diet and stable atorvastatin therapy; and (3) patients with other forms of primary hypercholesterolemia (not FH) with diet alone. A dose of 150 mg of the

anti-PCSK9 antibody subcutaneously on days 1, 29 and 43. Results of patient groups

who received the antibody (150-mg-FH-no; 150-mg-FH-Yes; 150-mg-combined) are shown in solid lines, while the results of patients who received a placebo (PBO-FH-no ; PBO-FH-Yes; PBO-combined) are shown in dashed lines.

Fig. 4 shows the study design of study 2 for the group of patients receiving a hypolipidemic treatment other than atorvastatin or not at a stable dose of atorvastatin 10 mg for at least 6 weeks before selection, or patients without prior drug. .

Fig. 5 shows the study design of study 2 for the group of patients receiving atorvastatin 10 mg at a stable dose for at least 6 weeks before selection.

Fig. 6 shows the distribution of the mean LDL-C values of patients in study 1 who receive 316P antibody in the treatment of stable atorvastatin for 12 weeks and LOCF (last data drag). The study was designed to evaluate the efficacy and safety of the 316P antibody in patients with hypercholesterolemia with elevated LDL-C (> 100 mg / dl or 2.59 mmol / l) treated with stable dose of atorvastatin (10 mg, 20 mg or 40 mg) During the preinclusion period, patients stabilized atorvastatin treatment (10 mg, 20 mg or 40 mg) if they were not yet. After an additional week, patients were randomized centrally by IVRS / IWRS in a 1: 1: 1: 1: 1: 1 ratio with respect to one of the 6 treatment groups (placebo, 316P 50 mg E2W, 316P 100 mg E2W, 316P 150 mg E2W, 316P 200 mg E4W, 316P 300 mg E4W) and were treated in a double blind form for approximately 12 weeks. Randomization is stratified by the dose of atorvastatin received before randomization. During the double-blind treatment period, patients returned to the site every 2 weeks to receive the study treatment (316P or placebo). The double-blind treatment period was then followed by a follow-up period of 8 weeks. As can be obtained from Figure 6, all treatment groups,

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Except for the group of patients who received placebo, they had a significant and persistent reduction in LDL-C levels with respect to the entire study period.

DETAILED DESCRIPTION OF THE INVENTION

Definitions

Before the present invention is described in detail below, it should be understood that the present invention is not limited to the particular methodology, protocols and reagents described herein, as these may vary. It should also be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to limit the scope of the present invention that will be limited only by the appended claims. Unless defined otherwise, all technical and scientific terms used herein have the same meanings that are commonly understood by a person skilled in the art to which the present invention belongs.

Preferably, the terms used herein are defined as described in "A multilingual glossary of biotechnological terms: (IUPAC Recommendations)", Leuenberger, H.G.W, Nagel, B. and Kolbl, H. eds. (1995), Helvetica Chimica Acta, CH-4010 Basel, Switzerland).

Throughout this specification and the claims that follow, unless the context requires otherwise, it will be understood that the word "understand", and variations such as "comprises" and "comprising", imply the inclusion of a number established integer or stage or group of integers or stages, but not the exclusion of any other integer or stage or group of integer or stage.

Several documents (for example: patents, patent applications, scientific publications, manufacturer specifications, instructions, sequence presentations with GenBank access number, etc.) are cited throughout the text of this specification. Nothing in this document should be construed as an admission that the invention has no right to precede such disclosure by virtue of the previous invention. In the event of a conflict between the definitions or teachings of such references and the incorporated definitions or teachings cited herein, the text of the present specification has priority.

Sequences: All sequences cited herein are disclosed in the attached sequence listing which, with its complete content and disclosure, is a part of this specification.

The term "approximately", when used with respect to a numerical value, is indicated to encompass numerical values within a range that has a lower limit that is 5% smaller than the indicated numerical value and that has an upper limit than It is 5% larger than the indicated numerical value.

The term "proprotein convertase subtilisin / human type 9 kexin" or "PCSK9h", as used herein, refers to PCSK9h having the nucleic acid sequence shown in SEQ ID NO: 754 and the amino acid sequence of SEQ ID NO: 755, or a biologically active fragment thereof.

The terms "specifically binds", "specific binding" or the like, mean that an antibody or antigen-binding fragment in the same way complexes with an antigen that is relatively stable under physiological conditions. Specific binding can be characterized by an equilibrium dissociation constant of at least about 1x10 "6 M or less (for example, a smaller Kd indicates a stronger bond). Methods for determining whether two molecules specifically bind are well known in the art and include, for example, equilibrium dialysis, surface plasmon resonance.An isolated antibody that specifically binds to PCSK9h may, however, exhibit cross-reactivity with other antigens such as PCSK9 molecules of other species. multi-specific (eg, bispecific) that bind PCSK9h and one or more additional antigens are considered, however, antibodies that "specifically bind" to PCSK9h, as used herein.

The term "Kd", as used herein, is intended to refer to the equilibrium dissociation constant of a particular antibody-antigen interaction. The equilibrium dissociation constant is usually measured in "mol / l" (abbreviated "M").

By the term "dissociation rate", "Kdis' or" kd "indicates an antibody that dissociates from PCSK9h with a rate constant of 1 x 10" 3 s "1 or less, preferably 1 x 10" 4 s " 1 or less, as determined by surface plasmon resonance, for example, BIACORE ™.

The term "high affinity" antibody refers to those mAbs that have a PCSK9h binding affinity of at

10 1 11 1 1 12

minus 10 'M; preferably 10 'M; even more preferably 10 'M, as measured by resonance of surface plasmons, for example, BIACORE ™ or ELISA of affinity in solution.

The term "surface plasmon resonance", as used herein, refers to an optical phenomenon that allows the analysis of biospecific interactions in real time by detection of alterations.

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at protein concentrations within a matrix of biosensors, for example, using the BIACORE ™ system (Pharmacia Biosensor AB, Uppsala, Sweden and Piscataway, N.J.).

An "epitope", also known as an antigenic determinant, is the region of an antigen that is recognized by the immune system, specifically by antibodies, B lymphocytes or T lymphocytes. As used herein, an "epitope" is the part of an antigen capable of binding to an antibody or antigen binding fragment thereof as described herein. In this context, the term "union" preferably refers to a "specific union", as defined herein. Epitopes normally consist of chemically active surface groups of molecules such as amino acids, sugar side chains, phosphoryl groups or sulfonyl groups, and may have specific three-dimensional structural characteristics and / or specific loading characteristics. Conformational and non-conformational epitopes can be distinguished in that the union to the former, but not to the latter, is lost in the presence of denaturing solvents.

A "paratope" is the part of an antibody that specifically binds to the epitope.

The term "antibody", as used herein, is intended to refer to immunoglobulin molecules comprising four polypeptide chains, two heavy chains (H) and two light chains (L), interconnected by disulfide bonds. The term "antibody" also includes all recombinant forms of antibodies, in particular of the antibodies described herein, for example, prokaryotic-expressed antibodies, non-glycosylar antibodies, and any fragment of antigen-binding antibody and derivatives as described. describe later. Each heavy chain is comprised of a variable region of the heavy chain ("HCVR" or "VH") and a constant region of the heavy chain (comprised of domains CH1, CH2 and CH3). Each light chain is comprised of a variable region of the light chain ("LCVR or" VL ") and a constant region of the light chain (CL). The VH and VL regions can be further subdivided into regions of hypervariability, called determining regions of Complementarity (CDR), interspersed with regions that are more conserved, called structural regions (FR) Each VH and VL is composed of three CDRs and four FRs, arranged from the amino end to the carboxy end in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4 The variable regions of the heavy and light chains contain a binding domain that interacts with an antigen.The constant regions of the antibodies can mediate the binding of immunoglobulin to tissues or host factors , which include various cells of the immune system (eg effector cells) and the first component (C1q) of the classical complement system.

It is also possible to replace one or more remaining CDRs or omit one or more CDRs. Antibodies have been described in the scientific literature in which one or two CDRs can be dispensed with for binding. Padlan et al. (1995 FASEB J. 9: 133-139) analyzed the contact regions between antibodies and their antigens, based on published crystalline structures, and concluded that only about one fifth to one third of the remains of CDRs are placed actually in contact with the antigen. Padlan also found many antibodies in which one or two CDRs did not have amino acids in contact with an antigen (see, therefore, Vajdos et al. 2002 J Mol Biol 320: 415-428).

CDR residues that do not contact the antigen can be identified based on previous studies (for example, H60-H65 residues in CDRH2 are not frequently required), from regions of Kabat CDRs that are outside the Chothia regions , by molecular modeling and / or empirically. If a CDRs or residue (s) thereof is omitted, it is usually substituted with an amino acid that occupies the corresponding position in another human antibody sequence or a consensus of such sequences. Positions for substitution within CDR and amino acids to be substituted can also be empirically selected. Empirical substitutions may be conservative or non-conservative substitutions.

The term "antigen binding fragment" of an antibody (or simply "binding portion"), as used herein, refers to one or more fragments of an antibody that retain the ability to specifically bind PCSK9h. It has been shown that the antigen binding function of an antibody can be performed by fragments of a full length antibody. Examples of binding fragments encompassed within the term "antigen binding fragment" of an antibody include (i) Fab fragments, monovalent fragments consisting of the VL, VH, CL and CH domains; (ii) F (ab ') 2 fragments, bivalent fragments comprising two Fab fragments linked by a disulfide bridge in the hinge region; (iii) Fd fragments consisting of the VH and CH domains; (iv) Fv fragments consisting of the VL and VH domains of a single arm of an antibody, (v) dAb fragments (Ward et al., (1989) Nature 341: 544-546), which consist of a VH domain; (vi) isolated complementarity determining regions (CDRs), and (vii) combinations of two or more isolated CDRs that can optionally be joined by a synthetic connector. In addition, although the two domains of the Fv fragment, VL and VH, are encoded by separate genes, they can be linked, using recombinant methods, by a synthetic linker that allows them to be prepared as a single chain protein in which the pair of VL regions and VH forms monovalent molecules (known as single chain Fv (scFv); see, for example, Bird et al. (1988) Science 242: 423-426; and Huston et al. (1988) Proc. Natl. Acad. Sci. USA 85: 5879-5883). Such single chain antibodies are also intended to be encompassed within the term "antigen binding fragment" of an antibody. A further example is a binding domain immunoglobulin fusion protein comprising (i) a binding domain polypeptide that is fused with a polypeptide of the immunoglobulin hinge region, (ii) a constant region of the heavy chain of the CH2 immunoglobulin fused with the region

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hinge, and (iii) a constant region of the heavy chain of the immunoglobulin CH3 fused with the constant region CH2. The binding domain polypeptide can be a variable region of the heavy chain or a variable region of the light chain. Immunoglobulin fusion proteins of the binding domain are further disclosed in US 2003/0118592 and US 2003/0133939. These antibody fragments are obtained using conventional techniques known to those skilled in the art, and the fragments are screened for utility in the same manner as intact antibodies. Additional examples of "antigen binding fragments" are called microantibodies, which are derived from individual CDRs. For example, Heap et al. describe a microantibody of 17 amino acid residues derived from the heavy chain CDR3 of an antibody directed against the envelope glycoprotein of HIV-1 gp120 (Heap CJ et al. (2005) J. Gen. Virol. 86: 1791- 1800). Other examples include small antibody mimetics comprising two or more CDR regions that are fused together, preferably related structural regions. A small antibody mimetic such that comprises VH CDR1 and VL cDR3 linked by the related VH FR2 has been described by Qiu et al. (Qiu X-Q, et al. (2007) Nature biotechnology 25 (8): 921-929).

Thus, the term "antibody or antigen binding fragment thereof," as used herein, refers to immunoglobulin molecules and immunologically active portions of immunoglobulin molecules, that is, molecules that contain a binding site to the antigen that bind immunospecifically to an antigen.

Antibodies and antigen-binding fragments thereof usable in the invention may be of any animal origin that includes birds and mammals. Preferably, the antibodies or fragments are of human, chimpanzee, rodent (eg, mouse, rat, guinea pig or rabbit), chicken, turkey, pig, sheep, goat, camel, cow, horse, donkey, cat or dog origin. It is particularly preferred that the antibodies are of human or murine origin. The antibodies disclosed herein also include chimeric molecules in which a constant region of antibody derived from one species, preferably human, is combined with the antigen binding site derived from another species, for example, mouse. In addition, the antibodies disclosed herein include humanized molecules in which the antigen binding sites of an antibody derived from a non-human species (eg, mouse) are combined with constant and structural regions of human origin.

As exemplified herein, the antibodies disclosed herein can be obtained directly from hybridomas expressing the antibody, or they can be cloned and recombinantly expressed in a host cell (eg, a CHO cell, or a lymphocytic cell). Additional examples of host cells are microorganisms, such as E. coli, and fungi, such as yeast. Alternatively, they can be recombinantly produced in a transgenic non-human animal or plant.

The term "chimeric antibody" refers to those antibodies in which a portion of each of the heavy and light chain amino acid sequences is homologous to corresponding sequences in antibodies derived from a particular species or belonging to a particular class, while that the remaining segment of the chain is homologous to corresponding sequences in another species or class. Normally, the variable region of both light and heavy chains mimics the variable regions of antibodies derived from one species of mammals, while the constant portions are homologous to antibody sequences derived from another. A clear advantage of such chimeric forms is that the variable region may conveniently derive from presently known sources using readily available B lymphocytes or hybridomas of non-human host organisms in combination with constant regions derived from, for example, human cell preparations. Although the variable region has the advantage of ease of preparation and the specificity is not affected by the source, the constant region that is human is less likely to elicit an immune response from a human subject when antibodies are injected than what the region would do. constant from a non-human source. However, the definition is not limited to this particular example.

The term "humanized antibody" refers to a molecule that has an antigen binding site that is substantially derived from an immunoglobulin of a non-human species, in which the remaining immunoglobulin structure of the molecule is based on the structure and / or sequence of a human immunoglobulin. The antigen binding site may either comprise complete variable domains fused into constant domains, or only the complementarity determining regions (CDR) grafted into appropriate structural regions in the variable domains. Antigen binding sites may be not mutated or modified by one or more amino acid substitutions, for example, modified to resemble human immunoglobulins more closely. Some forms of humanized antibodies preserve all CDR sequences (for example, a humanized mouse antibody containing the six CDRs of the mouse antibody). Other forms have one or more CDRs that are CDRs that are altered from the original antibody.

Different methods for humanizing antibodies are known to the expert, as reviewed by Almagro and Fransson (Almagro JC and Fransson J (2008) Frontiers in Bioscience 13: 1619-1633). Almagro and Fransson distinguish between rational and empirical approaches. Rational approaches are characterized by generating some variants of the modified antibody and evaluating its binding or any other property of interest. If the designed variants do not produce the expected results, a new cycle of design and evaluation of the joint is initiated. Rational approaches include CDR grafting, surface conditioning, superhumanization and optimization of human chain content. In contrast, empirical approaches are based on the generation of large

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libraries of humanized variants and selection of the best clones using high performance enrichment or screening technologies. Therefore, empirical approaches depend on a reliable selection and / or screening system that is capable of searching through a wide space of antibody variants. In vitro presentation technologies, such as phage display and ribosome presentation, meet these requirements and are well known to the expert. Empirical approaches include FR libraries, guided selection, shuffling of structural regions and Humaneering.

The term "human antibody", as used herein, is intended to include antibodies that have variable and constant regions derived from human germline immunoglobulin sequences. The human mAbs disclosed herein may include amino acid residues not encoded by human germline immunoglobulin sequences (eg, mutations introduced by random or site-specific mutagenesis in vitro or by in vivo somatic mutation), for example , in the CDRs and in particular CDR3. However, the term "human antibody", as used herein, is not intended to include mAbs in which CDR sequences derived from the germ line of other mammalian species (eg, mouse), have been grafted into sequences. of human FR. The human antibodies disclosed herein include antibodies isolated from human immunoglobulin libraries or from transgenic animals for one or more human immunoglobulins and which do not express endogenous immunoglobulins, as described, for example, in US Pat. No. 5,939,598 by Kucherlapati and Jakobovits.

The term "monoclonal antibody", as used herein, refers to a preparation of antibody molecules of unique molecular composition. A monoclonal antibody shows a unique binding specificity and affinity for a particular epitope. In one embodiment, the monoclonal antibodies are produced by a hybridoma that includes a B lymphocyte obtained from a non-human animal, for example, mouse, fused with an immortalized cell.

The term "recombinant antibody", as used herein, includes all antibodies that are prepared, expressed, created or isolated by recombinant means, such as (a) antibodies isolated from an animal (eg, a mouse) that is transgenic or transchromosomal with respect to immunoglobulin genes or a hybridoma prepared therefrom, (b) isolated antibodies from a transformed host cell to express the antibody, for example, from a transfectome, (c) isolated antibodies from a library of recombinant combinatorial antibodies, and (d) antibodies prepared, expressed, created or isolated by any other means involving the sequence splicing of immunoglobulin genes with other DNA sequences.

The term "transfectome," as used herein, includes recombinant eukaryotic host cells that express an antibody, such as CHO cells, NS / 0 cells, HEK293 cells, HEK293T cells, plant cells, or fungi, which include cells of yeast.

As used herein, a "heterologous antibody" is defined in relation to a transgenic organism that produces such an antibody. This term refers to an antibody that has an amino acid sequence or a coding nucleic acid sequence corresponding to that found in an organism that does not consist of the transgenic organism, and which is generally derived from a species other than the transgenic organism.

As used herein, a "heterohybrid antibody" refers to an antibody that has light and heavy chains of different organism origins. For example, an antibody that has a human heavy chain associated with a murine light chain is a heterohybrid antibody.

Thus, "antibodies and antigen binding fragments thereof" suitable for use in the present invention include, but are not limited to, polyclonal, monoclonal, monovalent, bispecific, heteroconjugate, multispecific, recombinant, heterologous, heterohybrid, chimeric antibodies. , humanized (in particular CDR grafted), deimmunized, or human, Fab fragments, Fab 'fragments, F (ab') 2 fragments, fragments produced by an expression library of Fab, Fd, Fv, disulfide-bound Fvs (dsFv ), single chain antibodies (eg scFv), diabodies or tetrabodies (Holliger P. et al. (1993) Proc. Natl. Acad. Sci. USA 90 (14), 6444-6448), nanobodies (also known as antibodies of single domain), anti-idiotypic (anti-Id) antibodies (including, for example, anti-Id antibodies for the antibodies disclosed herein) and epitope binding fragments of any of the foregoing.

The antibodies described herein are preferably isolated. An "isolated antibody", as used herein, is intended to refer to an antibody that is substantially free of other mAbs that have different antigenic specificities (eg, an isolated antibody that specifically binds to PCSK9h is substantially free of mAbs that specifically bind antigens other than PCSK9h). An isolated antibody that specifically binds to PCSK9h may, however, have cross-reactivity with other antigens, such as PCSK9 molecules from other species.

As used herein, a "PCSK9 antagonist" indicates a compound that inhibits at least one biological activity of PCSK9, preferably the proteinase activity of PCSK9. PCSK9 antagonists

Preferred are characterized in that they bind from 10% to 100% (preferably 50% to 100%) of PCSK9 present in the blood when used in stoichiometric amounts. Preferred PCSK9 antagonists of the present disclosure are neutralizing antibodies.

A "neutralizing antibody", as used herein (or an "antibody that neutralizes PCSK9 activity 5"), is intended to refer to an antibody whose binding to PCSK9h results in the inhibition of at least one biological activity of PCSK9, preferably inhibition of PCSK9 proteinase activity. This inhibition of the biological activity of PCSK9 can be evaluated by measuring one or more indicators of the biological activity of PCSK9 by one or more of several standard in vitro or in vivo assays known in the art. Such tests are described, for example, in US 2010/0166768 A1.

10 As PCSK9 increases LDL cholesterol in plasma by promoting degradation of the LDL receptor, the activity of PCSK9 has an effect on several diseases associated with high levels of LDL cholesterol in plasma. Therefore, PCSK9 antagonists, such as neutralizing anti-PCSK9h antibodies or antigen binding fragments thereof, are useful for reducing high total cholesterol, non-HDL cholesterol, LDL cholesterol and / or apolipoprotein B100 (ApoB100). Therefore, PCSK9 antagonists are useful for improving,

15 correct, inhibit or prevent several such diseases, including without limitation hypercholesterolemia, hyperlipidemia, dyslipidemia, atherosclerosis and cardiovascular diseases.

In specific embodiments, anti-PCSK9 antibodies or antigen binding fragments thereof described herein can be conjugated to a therapeutic moiety ("immunoconjugate"), such as a cytotoxin, a chemotherapeutic drug, an immunosuppressant or a radioisotope.

A "conservative amino acid substitution" is one in which an amino acid residue is substituted by another amino acid residue having a side chain (group R) with similar chemical properties (eg, filler or hydrophobia). In general, a conservative amino acid substitution will not substantially change the functional properties of a protein. In cases where two or more amino acid sequences are differentiated from each other by conservative substitutions, the percentage or degree of similarity can be adjusted high to correct the nature

25 conservative substitution. Means of preparing this adjustment are well known to those skilled in the art. See, for example, Pearson (1994) Methods Mol. Biol. 24: 307-331. Examples of amino acid groups having side chains with similar chemical properties include

1) aliphatic side chains: glycine, alanine, valine, leucine and isoleucine;

2) aliphatic hydroxyl side chains: serine and threonine;

30 3) side chains containing amide: asparagine and glutamine;

4) aromatic side chains: phenylalanine, tyrosine and tryptophan;

5) basic side chains: lysine, arginine and histidine;

6) acidic side chains: aspartate and glutamate, and

7) sulfur-containing side chains: cysteine and methionine.

Preferred conservative amino acid substitution groups are: valine-leucine-isoleucine, phenylalanine-tyrosine, lysine-arginine, alanine-valine, glutamate-aspartate and asparagine-glutamine. Alternatively, a conservative substitution is any change that has a positive value in the PAM250 logarithmic probability matrix disclosed in Gonnet et al. (1992) Science 256: 1443-45. A "moderately conservative" substitution is any change that has a non-negative value in the logarithmic probability matrix PAM250. Given the code

Known genetic, and recombinant and synthetic DNA techniques, the skilled scientist can easily construct DNAs encoding conservative amino acid variants.

As used herein, "non-conservative substitutions" or "non-conservative amino acid exchanges" are defined as exchanges of one amino acid for another amino acid listed in a different group of the seven standard amino acid groups 1) to 7) shown above. .

The term "substantial identity" or "substantially identical", when referring to a nucleic acid or fragment thereof, indicates that, when optimally aligned with appropriate nucleotide insertions or deletions with another nucleic acid (or its complementary strand), there is nucleotide sequence identity in at least about 90%, and more preferably at least about 95%, 96%, 97%, 98% or 99% of the nucleotide bases, as measured by any identity algorithm of well-known sequence, such as

50 FASTA, BLAST or GAP, as discussed below.

As applied to polypeptides, the term "substantial similarity" or "substantially similar" means that two peptide sequences, when optimally aligned, such as by the GAP or BESTFIT programs using default hole weights, share at least 90%. sequence identity, even more preferably at

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minus 95%, 98% or 99% sequence identity. Preferably, residue positions that are not identical are differentiated by conservative amino acid substitutions.

Sequence similarity for polypeptides is usually measured using sequence analysis software. Protein analysis software matches similar sequences using similarity measurements assigned to various substitutions, deletions and other modifications, including conservative amino acid substitutions. For example, GCG software contains programs such as GAP and BESTFIT that can be used with default parameters to determine sequence homology or sequence identity between closely related polypeptides, such as homologous polypeptides of different species of organisms or between a natural protein and a mutein of it. See, for example, GCG Version 6.1. Polypeptide sequences can also be compared using FASTA with default or recommended parameters; a program in gCg Version 6.1. FASTA (for example, FASTA2 and FASTA3) provides alignments and percent sequence identity of the regions of the best overlap between query and search sequences (Pearson (2000) above). Another preferred algorithm when comparing a sequence of the invention with a database containing a large number of sequences from different organisms is the BLAST software, especially BLASTP or TBLASTN, using default parameters. See, for example, Altschul et al. (1990) J. Mol. Biol. 215: 403 410 and (1997) Nucleic Acids Res. 25: 3389 402.

When referring to the sequence identity percentages in the present application, these percentages are calculated in relation to the full length of the longest sequence, if not specifically indicated otherwise. This calculation in relation to the full length of the longest sequence applies to both nucleic acid sequences and polypeptide sequences.

As used herein, "treat", "treat" or "treatment" of a disease or disorder means performing one or more of the following: (a) reduce the severity and / or duration of the disorder; (b) limit or prevent the development of symptoms characteristic of the disorder (s) being treated; (c) inhibit the worsening of symptoms characteristic of the disorder (s) being treated; (d) limit or prevent the recurrence of the disorder (s) in patients who have previously had the disorder (s); and (e) limit or prevent the recurrence of symptoms in patients who were previously symptomatic for the disorder (s).

As used herein, "prevent", "prevent", "prevention" or "prophylaxis" of a disease or disorder means preventing a disorder from occurring in the subject.

As used herein, the terms "is for administration" and "is for administration" have the same meaning as "preparing for administration". In other words, the declaration that an active compound "is for administration" has to be understood in that said active compound has been formulated and prepared in doses so that said active compound is in a state capable of exerting its therapeutic activity.

The terms "therapeutically effective amount" or "therapeutic amount" are intended to mean that the amount of a drug or pharmaceutical agent that will cause the biological or medical response of a tissue, a system, animal or human that is being sought by a researcher, veterinarian, medical doctor or other clinical professional. The term "prophylactically effective amount" is intended to mean that the amount of a pharmaceutical drug that will prevent or reduce the risk of occurrence of the biological or medical event that is sought to be prevented in a tissue, system, animal or human by a researcher, veterinarian, doctor doctor or other clinical professional. Particularly, the dosage that a patient receives can be selected to achieve the desired amount of LDL (low density lipoprotein) cholesterol reduction; The dosage that a patient receives can also be assessed over time in order to reach an objective LDL level. The dosage schedule that uses an antibody or an antigen binding fragment thereof as described herein is selected according to a variety of factors including type, species, age, weight, body mass index, sex and medical condition of the patient; the severity of the condition to be treated; the potency of the compound chosen to be administered; the route of administration; the end of administration; and renal and hepatic function of the patient.

As used herein, a "patient" means any human or non-human animal, such as a mammal, reptile or bird that can benefit from a treatment with the antibodies and antigen-binding fragments thereof described herein. . Preferably, a "patient" is selected from the group consisting of laboratory animals (eg, mouse or rat), domestic animals (including, for example, guinea pig, rabbit, chicken, turkey, pig, sheep, goat, camel, cow , horse, donkey, cat or dog), rodent or primates that include chimpanzee, gorilla, bonobo and humans. It is particularly preferred that the "patient" be a human being.

The terms "subject" or "individual" are used interchangeably in this document. As used herein, a "subject" refers to a human or non-human animal (for example, a mammal, avian, reptile, fish, amphibian or invertebrate; preferably an individual that can either benefit from one of the different aspects of the present disclosure (for example, a method of treatment or a drug identified by the present methods) or which can be used as a laboratory animal for the identification or characterization of a drug or a method of treatment. for example, being a human being, a wild animal, an animal

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domestic or laboratory animal; Examples include: mammal, for example, human being, non-human primate (chimpanzee, bonobo, gorilla), dog, cat, rodent (for example, mouse, guinea pig, rat, hamster or rabbit, horse, donkey, cow, sheep, goat , pig, camel; avian, such as duck, pigeon, turkey, goose or chick; reptile such as: turtle, galapago, snake, lizard, amphibian such as frog (eg, Xenopus laevis); fish such as koi or zebrafish ; invertebrate such as a worm (for example, C. elegans) or an insect (such as a fly, for example, Drosophila melanogaster.) The term "individual" also includes the different stages of morphological development of avian, fish, reptiles or insects, such as egg, pupa, larva or imago It is further preferred if the subject is a "patient".

As used herein, "unit dosage form" refers to physically discrete units suitable as unit dosages for human and / or animal subjects, each unit containing a predetermined amount of active material (eg, about 50 to about 500 mg of antibody against PCSK5 and / or, for example, 0.05 mg to 100 mg of HMG-CoA reductase inhibitor) calculated to produce the desired therapeutic effect in association with the required diluent, carrier or pharmaceutical carrier. The specifications for the novel unit dosage forms of the present disclosure are imposed by and are directly dependent on (a) the unique characteristics of the active material and the particular therapeutic effect to be achieved, and (b) the limitation inherent in the technique. of combining such an active material for therapeutic use in animals or humans, as disclosed in this specification, these characteristics being the present disclosure. Examples of suitable unit dosage forms according to the present disclosure are vials, tablets, capsules, troches, suppositories, powder sachets, wafers, seals, ampoules, segregated multiples of any of the foregoing, and other forms as described herein. or generally known in the art. One or more such unit dosage forms of the antibody may be comprised in an article of manufacture disclosed herein, which optionally further comprises one or more unit dosage forms of an HMG-CoA reductase inhibitor (eg, a container alveolate of tablets comprising as active ingredient the HMG-CoA reductase inhibitor).

The term "active material" refers to any material with therapeutic activity, such as one or more active ingredients. The active ingredients to be used as therapeutic agents can be easily prepared in such a unit dosage form with the use of pharmaceutical materials that are themselves available in the art and can be prepared by established procedures.

The following preparations are illustrative of the preparation of the unit dosage forms of the present disclosure, and not as a limitation thereof. Various dosage forms can be prepared that are part of this disclosure. For example, a unit dosage per vial may contain 0.5 ml, 1 ml, 2 ml, 3 ml, 4 ml, 5 ml, 6 ml, 7 ml, 8 ml, 9 ml, 10 ml, 15 ml or 20 ml of antibody against PCSK5 or a fragment thereof ranging from about 40 to about 500 mg of antibody against PCSK5. If necessary, these preparations can be adjusted to a desired concentration by adding a sterile diluent to each vial. In one embodiment, the formulation components of the disclosure are supplied either separately or mixed together in unit dosage form, for example, as a dry lyophilized powder or concentrated without water in a hermetically sealed container such as a vial, a vial or envelope that indicates the amount of active agent. Where the composition is to be administered by infusion, it can be dispensed with an infusion bottle containing sterile pharmaceutical grade water or saline solution. Where the composition is administered by injection, a sterile water vial for injection or saline solution can be provided so that the components can be mixed before administration.

Disclosure formulations include bulk drug compositions useful in the manufacture of pharmaceutical compositions (eg, compositions that are suitable for administration to a subject or patient) that can be used in the preparation of unit dosage forms. In a preferred embodiment, a composition of the disclosure is a pharmaceutical composition. Such compositions comprise a prophylactically or therapeutically effective amount of one or more prophylactic or therapeutic agents (for example, a disclosure antibody or other prophylactic or therapeutic agent), and a pharmaceutically acceptable carrier. Preferably, the pharmaceutical compositions are formulated to be suitable for the route of administration to a subject.

The active materials or components (for example, antibodies or fragments thereof and HMG-CoA reductase inhibitors) can be formulated as various dosage forms that include solid dosage forms for oral administration such as capsules, tablets, pills, powders. and granules, liquid dosage forms for oral administration such as pharmaceutically acceptable emulsions, microemulsions, solutions, suspensions, syrups and elixirs, injectable preparations, for example, sterile injectable aqueous or oleaginous suspensions, compositions for rectal or vaginal administration, preferably suppositories , and dosage forms for topical or transdermal administration such as ointments, pastes, creams, lotions, gels, powders, solutions, sprays, inhalants or patches.

In a specific embodiment, the term "pharmaceutically acceptable" means authorized by a US federal government regulatory agency. or state or AEM (European Medicines Agency) or listed in the US Pharmacopoeia. (United States Pharmacopeia-33 / National Formulary-28 Reissue, published by United States Pharmacopeia Convention, Inc., Rockville Md., Publication date: April 2010) or other pharmacopoeia

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generally recognized for use in animals, and more particularly in humans. The term "vehicle" refers to a diluent, adjuvant {for example, Freund's adjuvant (complete and incomplete)), excipient, or vehicle with which the therapeutic drug is administered. Such pharmaceutical vehicles may be sterile liquids, such as water and oils, which include those of petroleum, animal, vegetable or synthetic origin, such as peanut oil, soybean oil, mineral oil, sesame oil. Water is a preferred vehicle when the pharmaceutical composition is administered intravenously. Saline solutions and aqueous solutions of dextrose and glycerol can also be used as liquid carriers, particularly for injectable solutions. Suitable pharmaceutical excipients include starch, glucose, lactose, sucrose, gelatin, malt, rice, flour, limestone, silica gel, sodium stearate, glycerol monostearate, talc, sodium chloride, skimmed milk powder, glycerol, propylene, glycol, water, ethanol For the use of (additional) excipients and their use, see also "Handbook of Pharmaceutical Excipients", fifth edition, R.C. Rowe, P.J. Seskey and S.C. Owen, Pharmaceutical Press, London, Chicago. The composition, if desired, may also contain minor amounts of wetting or emulsifying agents, or pH buffering agents. These compositions can take the form of solutions, suspensions, emulsion, tablets, pills, capsules, powders, sustained release formulations. The oral formulation may include standard vehicles such as pharmaceutical grades of mannitol, lactose, starch, magnesium stearate, sodium saccharin, cellulose, magnesium carbonate, etc. Examples of suitable pharmaceutical vehicles are described in "Remington's Pharmaceutical Sciences" by E. W. Martin. Such compositions will contain a prophylactically or therapeutically effective amount of the antibody, preferably in purified form, together with a suitable amount of vehicle so as to provide the form for proper administration to the patient. The formulation must be adapted to the mode of administration.

Generally, the components of compositions of the disclosure are supplied either separately or mixed together in unit dosage form, for example, as a dry formulation for dissolution such as a lyophilized powder, freeze dried powder or concentrated without water in a container Hermetically sealed such as a blister or envelope that indicates the amount of active agent. The components of the disclosed compositions may also be supplied as a mixed liquid formulation (i.e. solution for injection or infusion) in a hermetically sealed container such as a vial, envelope, a pre-filled syringe or auto-injector, or a cartridge for a syringe or applicator reusable (for example, pen or auto-injector). Where the composition is to be administered by infusion, it can be dispensed with an infusion bottle containing sterile pharmaceutical grade water or saline solution. Where the composition is administered by injection, a sterile water vial for injection or saline solution can be provided so that the components can be mixed before administration.

The disclosure also provides that the formulation is packaged in a hermetically sealed container such as a blister or envelope indicating the amount of antibody. In one embodiment, the formulation disclosed herein comprising an antibody is supplied as a dry formulation, such as a sterilized lyophilized powder, freeze dried powder, spray dried powder or concentrated without water in a hermetically sealed container and can be reconstituted , for example, with water or saline solution at the appropriate concentration for administration to a subject. In another embodiment, the antibody or antigen binding fragment thereof is supplied as a liquid formulation such as a solution for injection or infusion. In one embodiment, the formulation disclosed herein comprising an antibody is supplied as a dry formulation or as a liquid formulation in a tightly sealed container at a unit dosage of at least 40 mg, at least 50 mg, more preferably at least 75 mg, at least 100 mg, at least 150 mg, at least 200 mg, at least 250 mg, at least 300 mg, at least 350 mg, at least 400 mg, at least 450 mg, or at least 500 mg, of antibody or antigen binding fragment thereof. The lyophilized formulation disclosed herein comprising an antibody should be stored at 2 to 8 ° C in its original container and the antibody should be administered within 12 hours, preferably within 6 hours, within 5 hours. hours, within 3 hours, or within 1 hour after being reconstituted. The formulation disclosed herein comprising antibodies can be formulated as neutral or salt forms. Pharmaceutically acceptable salts include those formed with anions such as those derived from hydrochloric, phosphoric, acetic, oxalic, tartaric acids, etc., and those formed with cations such as those derived from sodium, potassium, ammonium, calcium, ferric, isopropylamine hydroxides. , triethylamine, 2- ethylaminoethanol, histidine, procaine, etc.

Adult subjects are characterized by having "hypertension" or high blood pressure when they have a systolic blood pressure greater than 140 mmHg and / or a diastolic blood pressure greater than 90 mmHg.

Specific populations treatable by the therapeutic methods of the disclosure include subjects with one or more of the following conditions: subjects indicated for apheresis of LDL, subjects with activating mutations of PCSK9 (increased function mutations, "GOF"), subjects with elevated levels of total cholesterol, subjects with high levels of low-density lipoprotein cholesterol (LDL-C), subjects with primary hypercholesterolemia, such as primary subjects with familial or unfamiliar hypercholesterolemia, subjects with heterozygous familial hypercholesterolemia (heFH); subjects with hypercholesterolemia, especially primary hypercholesterolemia, who are intolerant to statins or uncontrolled with statins; and subjects at risk of developing hypercholesterolemia that can be treated preventively. Other indications include hyperlipidemia and dyslipidemia, especially if they are associated with secondary causes such as type 2 diabetes mellitus, cholestatic liver diseases (primary biliary cirrhosis), nephrotic syndrome, hypothyroidism, obesity; and the prevention and treatment of

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atherosclerosis and cardiovascular diseases, such as coronary heart disease (CHD). The conditions or disorders as listed for the above populations or subjects are conditions or disorders, for which it is disclosed that the treatment with the antibody herein is especially suitable.

However, depending on the severity of the aforementioned diseases and conditions, the treatment of the subjects with the antibodies and antigen binding fragments disclosed herein may be contraindicated for certain diseases and conditions.

The term "adverse effect" (or side effect) refers to a harmful and unwanted effect resulting from a medication. An adverse effect can be called a "side effect", when it is evaluated as being secondary to a main or therapeutic effect. Some adverse effects occur only when treatment is started, increased or interrupted. Adverse effects can cause medical complications of a disease and negatively affect its prognosis. Examples of side effects are allergic reactions, vomiting, headaches or dizziness or any other effect described herein.

As used herein, "treat", "treat" or "treatment" of a disease or disorder means performing one or more of the following: (a) reduce the severity and / or duration of the disorder; (b) limit or prevent the development of symptoms characteristic of the disorder (s) being treated; (c) inhibit the worsening of symptoms characteristic of the disorder (s) being treated; (d) limit or prevent the recurrence of the disorder (s) in patients who have previously had the disorder (s); and (e) limit or prevent the recurrence of symptoms in patients who were previously symptomatic for the disorder (s).

As used herein, "prevent", "prevent", "prevention" or "prophylaxis" of a disease, condition or disorder means preventing a disorder, disease or condition from occurring in the subject.

High levels of total cholesterol are understood in the context of the present disclosure to preferably be total cholesterol levels of 200 mg / dl or more, especially 240 mg / dl or more. International treatment guidelines recommend reducing LDL-C to <2.0-2.6 mmol / l (<77-100 mg / dl) in patients with established cardiovascular disease (CVD) and to <1.8-2.0 mmol / l (<70-77 mg / dl) in high-risk groups such as those with CVD plus diabetes, smoker, poorly controlled hypertension, metabolic syndrome or previous myocardial infarction. Elevated levels of LDL-C without being understood in the context of the present disclosure for being LDL-C levels of 77 mg / dL or more (especially for patients with one or more of the following characteristics: established CVD and one or more of [diabetes, with smoker, poorly controlled hypertension, metabolic syndrome or previous myocardial infarction]) and 100 mg / dl or more (especially for patients with established CVD), 130 mg / dl or more, or 160 mg / dl or 190 mg / dl or more. Low levels of high density lipoprotein (HDL levels) in the context of the present disclosure are understood to be preferably less than about 40 mg / dl.

The terms "not controlled by statins" or "statin resistant", especially in the context of hyperlipidemia, hypercholesterolemia etc., are used synonymously herein and refer to conditions such as hyperlipidemia, in which treatment with a statin (i.e., regular administration of a statin such as atorvastatin to a patient) does not significantly reduce total cholesterol or LDL-C or is not sufficient to establish normolipidemic levels for the patient or to establish a lipid level (for example, total cholesterol or LDL-C) which is not a significant risk factor for the development of cardiovascular diseases. This means, for example, that statin therapy is not sufficient to establish levels below 130 mg / dl in general, or below 100 mg / dl (for example, approximately> 77 mg / dl to approximately 100 mg / dl) , especially in patients with established cardiovascular diseases, or to establish levels of approximately less than 77 mg / dl (for example, approximately> 70-77 mg / dl), especially in high-risk groups such as those with CVD plus diabetes, smoker , poorly controlled hypertension, metabolic syndrome, or previous myocardial infarction. In the context of the present disclosure, statin resistance preferably refers to atorvastatin resistance.

Different aspects of the disclosure are defined in more detail in the following passages. Each aspect so defined can be combined with any other aspect or aspects, unless clearly stated otherwise. In particular, any characteristic indicated as being preferred or advantageous may be combined with any other characteristic or characteristics indicated as being preferred or advantageous, unless clearly stated otherwise.

In a first aspect, the present disclosure refers to a method of treatment of a disease or condition in which the expression or activity of PCSK9 causes a, comprising:

administering a therapeutic amount of an antibody or antigen-binding fragment thereof that specifically binds PCSK9h (proprotein convertase subtilisin / human type 9 kexin) to a subject in need thereof, in which the antibody or fragment binding to the antigen thereof is administered in a dose amount ranging from 5 mg to 500 mg, and

administering a therapeutic amount of an HMG-CoA reductase inhibitor to said subject, wherein the HMG-CoA reductase inhibitor is preferably administered in a dose amount ranging from 0.05 mg to 100 mg.

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In the context of the present application, the term "a disease or condition in which the expression or activity of PCSK9 causes an impact" is understood to mean any disease or condition in which the application of an antibody against PCSK-9 causes a impact.

In preferred embodiments of the first and other aspects of the present disclosure, the disease or condition in which the expression or activity of PCSK9 causes an impact is improved, corrected, inhibited or prevented with a PCSK9 antagonist.

In additional preferred embodiments of the first and other aspects of the present disclosure, the disease or condition is selected from the group consisting of: elevated levels of total cholesterol, elevated levels of low density lipoprotein cholesterol (C-LDL), hypercholesterolemia, particularly hypercholesterolemia not controlled by statins, hyperlipidemia, dyslipidemia, atherosclerosis, cardiovascular diseases, primary hypercholesterolemia, such as familial primary hypercholesterolemia or non-familial primary hypercholesterolemia (especially primary hypercholesterolemia) not controlled by statins (particularly not controlled by atorvastatin).

In preferred embodiments of the first and other aspects of the present disclosure, the subject in need thereof is a subject indicated for apheresis of LDL, a subject with activating mutations of PCSK9, a subject with heterozygous familial hypercholesterolemia, a subject with primary hypercholesterolemia, a subject with primary hypercholesterolemia that is not controlled with statins, a subject at risk of developing hypercholesterolemia, a subject with hypercholesterolemia, a subject with hyperlipidemia, a subject with dyslipidemia, a subject with atherosclerosis or a subject with cardiovascular disease. Most preferably, the subject in need thereof is a human subject.

In some embodiments of the first and other aspects of the disclosure, the HMG-CoA reductase inhibitor is administered three times per day, twice per day, or once per day. In some embodiments of the first and other aspects disclosed herein, the HMG-CoA reductase inhibitor is administered every day, every two days, every third day, every fourth day, every fifth day, or every sixth day. In some embodiments of the first and other aspects of the present disclosure, the HMG-CoA reductase inhibitor is administered every week, every two weeks, every three weeks, or every four weeks. In some embodiments of the first and other aspects of the present disclosure, the HMG-CoA reductase inhibitor is administered in the morning, at noon or in the afternoon. In preferred embodiments, the HMG-CoA reductase inhibitor is administered once a day, preferably orally, preferably in the afternoon.

Preferably, the HMG-CoA reductase inhibitor is a statin. More preferably, the statin is selected from the group consisting of cerivastatin, atorvastatin, simvastatin, pitavastatin, rosuvastatin, fluvastatin, lovastatin and pravastatin.

In more preferred embodiments of the first and other aspects of the present disclosure, the statin is

- cerivastatin administered at a daily dose of between 0.05 mg and 2 mg, preferably at a daily dose of 0.2 mg, 0.4 mg, or 0.8 mg;

- atorvastatin administered in a daily dose between 2 mg and 100 mg, preferably in a daily dose of 10 mg, 20 mg, 40 mg, or 80 mg;

- simvastatin administered in a daily dose between 2 mg and 100 mg, preferably in a daily dose of 5 mg, 10 mg, 20 mg, 40 mg, or 80 mg;

- pitavastatin administered in a daily dose between 0.2 mg and 100 mg, preferably in a daily dose of 1 mg, 2 mg, 5 mg, 10 mg, or 20 mg;

- rosuvastatin administered in a daily dose between 2 mg and 100 mg, preferably in a daily dose of 5 mg, 10 mg, 20 mg, or 40 mg;

- fluvastatin administered in a daily dose between 2 mg and 100 mg, preferably in a daily dose of 20 mg, 40 mg, or 80 mg;

- lovastatin administered at a daily dose between 2 mg and 100 mg, preferably at a daily dose of 10 mg, 20 mg, 40 mg, or 80 mg; or

- pravastatin administered in a daily dose between 2 mg and 100 mg, preferably in a daily dose of 10 mg, 20 mg, 40 mg, or 80 mg.

In preferred embodiments of the first and other aspects of the present disclosure, the antibody or antigen-binding fragment thereof is administered to the subject every two weeks, every four weeks or once a month. Administration is preferred every four weeks or administration once a month (that is, once per calendar month, for example, every first, second, etc. day of the month or every first, second, third Monday, Tuesday, etc. , every month, unlike administration every four weeks) in view of patient compliance. Be

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he prefers administration every two weeks in view of a very low variation in blood cholesterol levels. Other suitable time programs for administration of the antibody or antigen binding fragment thereof include without limitation one administration once a day, every two days, every third day, every fourth day, every fifth day, every sixth day, every week, every three weeks, every fifth week, every sixth week, every eighth week, every tenth week, and every twelfth week.

In preferred embodiments of the first and other aspects of the present disclosure, the antibody or antigen-binding fragment thereof is administered in a dose amount ranging for example, from about 40 mg to about 500 mg, from about 50 mg to about 500 mg, about 50 mg to 300 mg or about 100 mg to 200 mg. In more preferred embodiments, the antibody or antigen binding fragment thereof is administered in a dose amount of about 50 mg, about 100 mg, about 150 mg, about 200 mg, about 250 mg, about 300 mg, about 350 mg, about 400 mg, about 450 mg or about 500 mg. Doses of approximately 50 to approximately 200 mg, for example, approximately 50 mg, approximately 100 mg, approximately 150 mg or approximately 200 mg are especially suitable for a biweekly dosage schedule (i.e., administration every two weeks), doses of approximately 150 mg to about 400 mg, for example, about 150 mg, about 200 mg, about 250 mg, about 300 mg about 350 mg or about 400 mg are especially suitable for an administration schedule with longer intervals, for example, an administration every third or every four weeks or once a month.

Antibodies and antigen-binding fragments thereof that can be used to practice the first and other aspects of the present disclosure are described in the "Preferred antibodies" section.

In a second aspect, the present disclosure relates to an antibody or an antigen-binding fragment thereof that specifically binds PCSK9h (proprotein convertase subtilisin / human type 9 kexin) for use in the treatment of a disease or condition in which the expression or activity of PCSK9 causes an impact,

wherein the antibody or antigen binding fragment thereof is for administration in an amount of

dose ranging from 5 mg to 500 mg,

wherein the antibody or antigen binding fragment thereof is also for administration in

combination with an HMG-CoA reductase inhibitor at a dose amount ranging from 0.05 mg to 100 mg.

In preferred embodiments of the second and other aspects of the present disclosure, the disease or condition in which the expression or activity of PCSK9 causes an impact is improved, corrected, inhibited or prevented with a PCSK9 antagonist.

In additional preferred embodiments of the second and other aspects of the present disclosure, the disease or condition is selected from the group consisting of: elevated levels of low density lipoprotein cholesterol (LDL-C), hypercholesterolemia, particularly hypercholesterolemia not controlled by statins , hyperlipidemia, dyslipidemia, atherosclerosis and cardiovascular diseases, particularly primary hypercholesterolemia such as familial primary hypercholesterolemia or nonfamilial primary hypercholesterolemia.

In preferred embodiments of the second and other aspects of the present disclosure, the antibody or antigen-binding fragment thereof is for administration to a subject indicated for LDL apheresis, a subject with PCSK9 activating mutations, a subject with heterozygous familial hypercholesterolemia. , a subject with primary hypercholesterolemia, a subject with primary hypercholesterolemia that is not controlled with statins, a subject at risk of developing hypercholesterolemia, a subject with hypercholesterolemia, a subject with hyperlipidemia, a subject with dyslipidemia, a subject with atherosclerosis or a subject with cardiovascular diseases. Most preferably, the subject is a human subject.

In some embodiments of the second and other aspects of the present disclosure, the antibody or antigen-binding fragment thereof is for administration in combination with an HMG-CoA reductase inhibitor, which is to be administered three times per day, twice per day. day, or once a day. In some embodiments of the second and other aspects disclosed herein, the HMG-CoA reductase inhibitor is to be administered every day, every other day, every third day, every fourth day, every fifth day, or every sixth day. In some embodiments of the second and the other aspects disclosed herein, the HMG-CoA reductase inhibitor will be administered every week, every two weeks, every three weeks, or every four weeks. In some embodiments of the second and the other aspects disclosed herein, the HMG-CoA reductase inhibitor will be administered in the morning, at noon or in the afternoon. In preferred embodiments, the HMG-CoA reductase inhibitor is to be administered once a day, preferably orally, preferably in the afternoon.

In preferred embodiments of the second and other aspects of the present disclosure, the HMG-CoA reductase inhibitor is a statin. More preferably, the statin is selected from the group consisting of cerivastatin, atorvastatin, simvastatin, pitavastatin, rosuvastatin, fluvastatin, lovastatin and pravastatin.

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In more preferred embodiments of the second and other aspects of the present disclosure, the statin is

- cerivastatin to be administered at a daily dose of between 0.05 mg and 2 mg, preferably at a daily dose of 0.2 mg, 0.4 mg, or 0.8 mg;

- atorvastatin to be administered in a daily dose between 2 mg and 100 mg, preferably in a daily dose of 10 mg, 20 mg, 40 mg, or 80 mg;

- simvastatin to be administered in a daily dose between 2 mg and 100 mg, preferably in a daily dose of 5 mg, 10 mg, 20 mg, 40 mg, or 80 mg;

- pitavastatin to be administered in a daily dose between 0.2 mg and 100 mg, preferably in a daily dose of 1 mg, 2 mg, 5 mg, 10 mg, or 20 mg;

- rosuvastatin to be administered at a daily dose between 2 mg and 100 mg, preferably at a daily dose of 5 mg, 10 mg, 20 mg, or 40 mg;

- fluvastatin to be administered in a daily dose between 2 mg and 100 mg, preferably in a daily dose of 20 mg, 40 mg, or 80 mg;

- lovastatin to be administered in a daily dose between 2 mg and 100 mg, preferably in a daily dose of 10 mg, 20 mg, 40 mg, or 80 mg; or

- pravastatin to be administered in a daily dose between 2 mg and 100 mg, preferably in a daily dose of 10 mg, 20 mg, 40 mg, or 80 mg.

In preferred embodiments of the second and other aspects of the present disclosure, the antibody or antigen-binding fragment thereof is for administration to the subject every two weeks, every four weeks or once a month. Administration every four weeks or administration once a month is preferred in view of patient compliance. Administration every two weeks is preferred in view of a very low variation in blood cholesterol levels. Other suitable time programs for the administration of the antibody or antigen binding fragment thereof include, without limitation, one administration once a day, every two days, every third day, every fourth day, every fifth day, every sixth day, every week, every three weeks, every fifth week, every sixth week, every eighth week, every tenth week, and every twelfth week.

In preferred embodiments of the second and the other aspects of the present disclosure, the antibody or antigen binding fragment thereof is for administration in a dose amount ranging from about 40 mg to about 500 mg or from about 50 mg to about 500 mg or from about 50 mg to about 400 mg or from about 50 mg to about 300 mg, or from about 100 mg to about 300 mg or from about 100 mg to about 200 mg. In more preferred embodiments, the antibody or antigen binding fragment thereof is for administration in a dose amount of about 50 mg, about 100 mg, about 150 mg, about 200 mg, about 250 mg, about 300 mg, about 350 or about 400 mg.

In preferred embodiments of the second and other aspects of the present disclosure, the antibody or antigen-binding fragment thereof is for administration in a dose amount (i.e., a dosage schedule) ranging from about 50 mg to about 200 mg. every two weeks (E2W), preferably about 50 mg E2W, about 100 mg E2W, about 150 mg E2W, about 200 mg E2W, about 250 mg E2W or about 300 mg E2W, with about 50 mg E2W, about 100 mg even more preferred E2W, approximately 150 mg E2W, approximately 200 mg E2W. According to an especially advantageous embodiment of the second and the other aspects disclosed herein, the antibody or antigen-binding fragment thereof is for administration in an amount of dose (i.e., a dosage schedule) E2W of about 50 mg to about 200 mg, from about 100 mg to about 180 mg, from about 130 mg to about 170 mg, from about 140 to about 160 mg or about 90, about 100, about 110, about 120, about 130, about 140, about 145, about 150, about 155, about 160, about 170, about 180, about 190 or about 200 mg E2W, the dosage guidelines from about 145 mg to about 155 mg E2W, and particularly about 150 mg E2W, belonging to particularly preferred embodiments.

In other preferred embodiments of the second and other aspects of the present disclosure, the antibody or antigen binding fragment thereof is for administration in a dose amount ranging from about 100 mg to about 400 mg every four weeks (E4W), preferably about 100 mg E4W, about 150 mg E4W, about 200 mg E4W, about 250 mg E4W, about 300 mg E4W, about 350 mg E4W or about 400 mg E4W, dose amounts of about 190 to about 310 E4W, of about

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200 to about 300 mg E4W, about 190 to about 210 E4W, about 195 to about 205 E4W, about 290 to about 310 E4W, about 295 to about 305 E4W, about 200 mg E4W or about 300 mg E4W at particularly preferred embodiments. These dose amounts indicated for administration E4W are also suitable for administration once a month.

Antibodies and antigen-binding fragments thereof that can be used to practice the second and other aspects of the present disclosure are described in the "Preferred antibodies" section.

In a third aspect, the present disclosure refers to an article of manufacture comprising: (a) a packaging material; (b) an antibody or an antigen binding fragment thereof that specifically binds to PCSK9h; and (c) a label or leaflet contained within the packaging material that indicates that patients receiving treatment with said antibody or antigen binding fragment can be treated for a disease or condition selected from the group consisting of hypercholesterolemia, hyperlipidemia, dyslipidemia, atherosclerosis and cardiovascular diseases.

In a fourth aspect, the present disclosure refers to an article of manufacture comprising: (a) a packaging material; (b) an antibody or an antigen binding fragment thereof that specifically binds to PCSK9h; and (c) a label or leaflet contained within the packaging material that indicates the treatment of patients with said antibody or antigen binding fragment thereof together with the administration of an HMG-CoAtal inhibitor such as a statin.

In a fifth aspect, the present disclosure refers to an article of manufacture comprising (a) a packaging material; (b) an antibody or an antigen binding fragment thereof that specifically binds to PCSK9h; and (c) a label or leaflet indicating that the treatment of patients with said antibody or antigen binding fragment thereof together with an HMG-CoAtal inhibitor such as a statin is contraindicated for patients belonging to one or more of the following groups: (i) smokers; (ii) persons 70 years of age or older;

(iii) people suffering from hypertension; (iv) women who are pregnant; (v) women who are trying to get pregnant; (vi) women who are breastfeeding; (vii) people who have or have ever had a disease that affects the liver; (viii) people who had some unexplained abnormal blood test for liver function; (ix) people who drink excessive amounts of alcohol; (x) people who have kidney problems; (xi) people suffering from hypothyroidism; (xii) people suffering from muscular disorders; (xiii) people who have had previous muscle problems during treatment with lipid lowering medicine; (xiv) people who have serious problems with their breathing; (xv) people taking one or more of the following medicines: medicines that alter the way the immune systems work (for example, cyclosporine or antihistamines), antibiotics or antifungal medicines (for example, erythromycin, clarithromycin, ketoconazole, itraconazole , rifampicin, fusidic acid), medicines that regulate lipid levels (for example, gemfibrozil, colestipol), calcium channel blockers (for example, verapamil, diltiazem), medicines that regulate heart rhythm (digoxin, amiodarone), protease inhibitors used in the treatment of HIV (for example, nelfinavir), warfarin, oral contraceptives, antacids or St. John's wort; or (xvi) people who drink more than 0.1 l of grapefruit juice per day or who eat more than half a grapefruit per day; (xvii) people who have a body mass index (BMI) greater than 40; (xviii) people who have a body mass index (BMI) of less than 18; (xix) people suffering from type 1 diabetes or type 2 diabetes; (xx) people positive for hepatitis B or hepatitis C; (xxi) people who have a known sensitivity to therapeutic drugs of monoclonal antibodies; (xxii) people who have a neutrophil concentration less than 1500 / mm3; (xxiii) people who have a platelet concentration of less than 100,000 / mm3; (xxiv) males who have a serum creatinine level greater than 1.5 x ULN (upper limit of normal); (xxv) women who have a serum creatinine level greater than 1.4 x ULN (upper limit of normal); (xxvi) people who have a level of alanine transaminase (ALT) or level of aspartate transaminase (AST) greater than 2 x ULN; or (xxvii) people who have a CPK level greater than 3 x ULN.

In preferred embodiments of the third, fourth and fifth aspect, the antibody or antigen binding fragment is an antibody or antigen binding fragment as specified below in the "Preferred antibodies" section.

The label or leaflet according to the different aspects and embodiments of the disclosure, particularly with respect to the different articles of manufacture disclosed herein, can be any type of data carrier suitable for being disposed within the container or container or on the outside of the container or container Preferably, the data carriers (ie, label or, chip, bar code or pamphlet or label comprising a bar code, etc.) comprise information such as

(i) composition, formulation, concentration and total amount, identity of the active principle (s)

content (s) in the article of manufacture, that is, of the antibody or antigen fragments, HMG-inhibitor

CoA reductase, pharmaceutical composition, unit dosage form or formulation disclosed in the

present document

(ii) number and composition of the unit dosage form contained in the article of manufacture

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(iii) indications, contraindications of the antibody or antigen fragments, pharmaceutical composition, unit dosage form or formulation disclosed herein

(iv) (ii) subjects / patients or populations of subjects / patients indicated or contraindicated for treatment with the antibody or antigen fragments, pharmaceutical composition, unit dosage form or formulation disclosed herein.

(v) instructions for use, dosage guidelines and / or administration guidelines

(vi) quality information such as information on the starting / batch number of the antibody or antigen fragments, pharmaceutical composition, unit dosage form or formulation of the present invention, the manufacturing or assembly site or the expiration date or the deadline selling,

(vii) information regarding the correct storage or handling of the article of manufacture, of the device for administration, or of the antibody or antigen fragments, pharmaceutical composition, unit dosage form or formulation disclosed herein,

(iv) information regarding the composition of the buffer (buffers), diluent (s), reagent (s), excipients, carriers, antibody formulations or antigen fragments, pharmaceutical composition, unit dosage form or disclosed formulation in the present document,

(vi) a warning regarding possible consequences when an inadequate dosage or administration and / or use guidelines is given in contraindicated indications of patient populations.

In preferred embodiments of the third, fourth and fifth aspects, the label or leaflet contains reference to a method of medical treatment or use according to the first or second aspect and the embodiments of the first or second aspect as described herein.

A further preferred embodiment of the present disclosure combines the characteristics of the third aspect and the fourth aspect as described herein.

A further preferred embodiment of the present disclosure combines the characteristics of the third aspect and the fifth aspect as described herein.

A further preferred embodiment of the present disclosure combines the characteristics of the fourth aspect and the fifth aspect as described herein.

A further preferred embodiment of the present disclosure combines the characteristics of the third aspect, the fourth aspect and the fifth aspect as described herein.

In a sixth aspect, the present disclosure relates to a method of testing the efficacy of an antibody or an antigen binding fragment thereof that specifically binds to PCSK9h for the treatment of a disease or condition selected from the group consisting of hypercholesterolemia, hyperlipidemia, dyslipidemia, atherosclerosis and cardiovascular diseases or any of the other conditions or diseases according to the first or second aspect disclosed herein, said method comprising:

treating a selected patient population with said antibody or antigen binding fragment thereof, in which each patient in said population has a cholesterol level of (C-LDL) greater than 100 mg / dl; Y

determining the efficacy of said antibody or antigen binding fragment thereof by determining the level of LDL-C in the patient population before and after administration of said antibody or antigen binding fragment thereof, in which a reduction in the LDL-C level of at least 25% with respect to a predose level in at least 75% of the patient population indicates that said antibody or antigen-binding fragment thereof is effective for the treatment of said disease or condition in said patient population.

In a seventh aspect, the present disclosure relates to a method of testing the efficacy of an antibody or an antigen binding fragment thereof that specifically binds to PCSK9h for the treatment of a disease or condition selected from the group consisting of hypercholesterolemia, hyperlipidemia, dyslipidemia, atherosclerosis and cardiovascular diseases or any of the other conditions or diseases according to the first or second aspect disclosed herein, said method comprising:

determining the efficacy of an antibody or antigen binding fragment thereof that has been used for the treatment of a selected patient population with said antibody or antigen binding fragment thereof, in which each patient in said population has a level of cholesterol of (C-LD) L higher than 100 mg / dl determining the level of C-LDL in the patient population before and after the administration of said antibody or antigen-binding fragment thereof, in which a reduction of the LDL-C level of at least 25% with respect to a predose level in at least 75% of the patient population indicates

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that said antibody or antigen binding fragment thereof is effective for the treatment of said disease or condition in said patient population.

In preferred embodiments of the sixth and seventh aspect, each patient in said population has received a lipid-lowering treatment by administration of an HMG-CoA inhibitor, such as a statin for at least 6 weeks before treatment with said antibody or antigen-binding fragment. of the same.

In preferred embodiments of the sixth and seventh aspect, the antibody or antigen binding fragment is an antibody or antigen binding fragment as specified below in the "Preferred antibodies" section.

In preferred embodiments of the sixth and seventh aspect, the selected patient population is treated with a treatment method according to the first aspect and the embodiments of the first or second aspect as described herein.

In an eighth aspect, the present disclosure relates to a package comprising an antibody or antigen binding fragment thereof that specifically binds PCSK9h and a label, said label comprising a printed statement informing the patient that the treatment of the Antibody together with an HMG-CoA reductase inhibitor such as a statin is indicated in one or more of the indications selected from the group consisting of hypercholesterolemia, hyperlipidemia, dyslipidemia, atherosclerosis and cardiovascular diseases or any of the other conditions or diseases according to the first or second aspect disclosed in this document. Antibodies and antigen-binding fragments thereof that can be used to practice the eighth aspect of the present disclosure are described in the "Preferred antibodies" section.

In a ninth aspect, the present disclosure refers to a package comprising an antibody or antigen-binding fragment thereof that specifically binds to PCSK9h and a label, said label comprising a printed statement informing the patient that the treatment of the Antibody together with a statin is contraindicated for patients belonging to one or more of the following groups: (i) smokers; (ii) persons 70 years of age or older; (iii) people suffering from hypertension; (iv) women who are pregnant; (v) women who are trying to get pregnant; (vi) women who are breastfeeding; (vii) people who have or have ever had a disease that affects the liver; (viii) people who had some unexplained abnormal blood test for liver function; (ix) people who drink excessive amounts of alcohol; (x) people who have kidney problems; (xi) people suffering from hypothyroidism; (xii) people suffering from muscular disorders; (xiii) people who have had previous muscle problems during treatment with lipid lowering medicine; (xiv) people who have serious problems with their breathing; (xv) people taking one or more of the following medicines: medicines that alter the way the immune systems work (for example, cyclosporine or antihistamines), antibiotics or antifungal medicines (for example, erythromycin, clarithromycin, ketoconazole, itraconazole , rifampicin, fusidic acid), medicines that regulate lipid levels (for example, gemfibrozil, colestipol), calcium channel blockers (for example, verapamil, diltiazem), medicines that regulate heart rhythm (digoxin, amiodarone), protease inhibitors used in the treatment of HIV (for example, nelfinavir), warfarin, oral contraceptives, antacids or St. John's wort; or (xvi) people who drink more than 0.1 l of grapefruit juice per day or who eat more than half a grapefruit per day; (xvii) people who have a body mass index (BMI) greater than 40; (xviii) people who have a body mass index (BMI) of less than 18; (xix) people suffering from type 1 diabetes or type 2 diabetes; (xx) people positive for hepatitis B or hepatitis C; (xxi) people who have a known sensitivity to therapeutic drugs of monoclonal antibodies; (xxii) people who have a neutrophil concentration less than 1500 / mm3; (xxiii) people who have a platelet concentration of less than 100,000 / mm3; (xxiv) males who have a serum creatinine level greater than 1.5 x ULN (upper limit of normal); (xxv) women who have a serum creatinine level greater than 1.4 x ULN (upper limit of normal); (xxvi) people who have a level of alanine transaminase (ALT) or level of aspartate transaminase (AST) greater than 2 x ULN; or (xxvii) people who have a CPK level greater than 3 x ULN. Antibodies and antigen-binding fragments thereof that can be used to practice the ninth aspect of the present disclosure are described in the "Preferred antibodies" section.

A further preferred embodiment of the present disclosure combines the characteristics of the eighth aspect and the ninth aspect as described herein.

In a tenth aspect, the present disclosure relates to a method of regulating the level of LDL in the blood comprising:

administering a therapeutic amount of an antibody or antigen-binding fragment thereof that specifically binds PCSK9h (proprotein convertase subtilisin / human type 9 kexin) to a subject in need thereof, in which the antibody or fragment binding to the antigen thereof is administered in a dose amount ranging from 5 mg to 500 mg, and

administering a therapeutic amount of an HMG-CoA reductase inhibitor to said subject, wherein the HMG-CoA reductase inhibitor is preferably administered in a dose amount ranging from 0.05 mg to 100 mg.

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In a eleventh aspect, the present disclosure relates to a method of preventing effects of a (persistently) high level of LDL in the blood comprising:

administering a therapeutic amount of an antibody or antigen-binding fragment thereof that specifically binds PCSK9h (proprotein convertase subtilisin / human type 9 kexin) to a subject in need thereof, in which the antibody or fragment binding to the antigen thereof is administered in a dose amount ranging from 5 mg to 500 mg, and

administering a therapeutic amount of an HMG-CoA reductase inhibitor to said subject, wherein the HMG-CoA reductase inhibitor is preferably administered in a dose amount ranging from 0.05 mg to 100 mg.

In preferred embodiments of the eleventh and eleventh aspects, the disease or condition in which the expression or activity of PCSK9 causes an impact is improved, corrected, inhibited or prevented with a PCSK9 antagonist. In additional preferred embodiments of the eleventh and eleventh aspect, the disease or condition in which the expression or activity of PCSK9 causes an impact is selected from the group consisting of: elevated levels of LDL-C, hypercholesterolemia, hyperlipidemia, dyslipidemia, atherosclerosis and cardiovascular diseases or any of the other conditions or diseases according to the first or second aspect disclosed herein.

In preferred embodiments of the eleventh and eleventh aspect, the subject in need thereof is a subject indicated for apheresis of LDL, a subject with activating mutations of PCSK9, a subject with heterozygous familial hypercholesterolemia, a subject with primary hypercholesterolemia that is not controlled with statins, a subject at risk of developing hypercholesterolemia, a subject with hypercholesterolemia, a subject with hyperlipidemia, a subject with dyslipidemia, a subject with atherosclerosis or a subject with cardiovascular disease or any of the subjects as described in the first or second aspect revealed in the present document. Most preferably, the subject in need thereof is a human subject.

In some embodiments of the eleventh and eleventh aspect, the HMG-CoA reductase inhibitor is administered three times per day, twice per day, or once per day. In some embodiments, the HMG-CoA reductase inhibitor is administered every day, every two days, every third day, every fourth day, every fifth day, or every sixth day. In some embodiments, the HMG-CoA reductase inhibitor is administered every week, every two weeks, every three weeks, every four weeks, or every month. In some embodiments, the HMG-CoA reductase inhibitor is administered in the morning, at noon or in the afternoon. In preferred embodiments, the HMG-CoA reductase inhibitor is administered once a day, preferably orally, preferably in the afternoon. Additional suitable administration guidelines are described in the first or second aspect.

Preferably, the HMG-CoA reductase inhibitor is a statin. More preferably, the statin is selected from the group consisting of cerivastatin, atorvastatin, simvastatin, pitavastatin, rosuvastatin, fluvastatin, lovastatin and pravastatin.

In more preferred embodiments of the eleventh and eleventh aspect, the statin is

- cerivastatin administered at a daily dose of between 0.05 mg and 2 mg, preferably at a daily dose of 0.2 mg, 0.4 mg, or 0.8 mg;

- atorvastatin administered in a daily dose between 2 mg and 100 mg, preferably in a daily dose of 10 mg, 20 mg, 40 mg, or 80 mg;

- simvastatin administered in a daily dose between 2 mg and 100 mg, preferably in a daily dose of 5 mg, 10 mg, 20 mg, 40 mg, or 80 mg;

- pitavastatin administered in a daily dose between 0.2 mg and 100 mg, preferably in a daily dose of 1 mg, 2 mg, 5 mg, 10 mg, or 20 mg;

- rosuvastatin administered in a daily dose between 2 mg and 100 mg, preferably in a daily dose of 5 mg, 10 mg, 20 mg, or 40 mg;

- fluvastatin administered in a daily dose between 2 mg and 100 mg, preferably in a daily dose of 20 mg, 40 mg, or 80 mg;

- lovastatin administered at a daily dose between 2 mg and 100 mg, preferably at a daily dose of 10 mg, 20 mg, 40 mg, or 80 mg; or

- pravastatin administered in a daily dose between 2 mg and 100 mg, preferably in a daily dose of 10 mg, 20 mg, 40 mg, or 80 mg.

In preferred embodiments of the tenth and eleventh aspect, the antibody or antigen binding fragment thereof is administered to the subject every two weeks, every four weeks or once a month. The

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administration every four weeks or every month in view of patient compliance. Administration every two weeks is preferred in view of a very low variation in blood cholesterol levels. Other suitable time programs for administration of the antibody or antigen-binding fragment thereof include, without limitation, administration once a day, every two days, every third day, every fourth day, every fifth day, every sixth day, every week, every three weeks, every fifth week, every sixth week, every eighth week, every tenth week, and every twelfth week.

In preferred embodiments of the eleventh and eleventh aspect, the antibody or antigen binding fragment thereof is administered in a dose amount ranging from 50 mg to 300 mg, for example, from 100 mg to 200 mg. In more preferred embodiments, the antibody or antigen binding fragment thereof is administered in a dose amount of about 50 mg, about 100 mg, about 150 mg, about 200 mg, or about 300 mg. Additional suitable and preferred dosage guidelines are described in the first or second aspect.

Antibodies and antigen-binding fragments thereof that can be used to practice the eleventh and eleventh aspects of the present disclosure are described in the "Preferred antibodies" section.

In a twelfth aspect, the present disclosure relates to a method of determining whether a pharmaceutical compound is usable to improve, correct, inhibit or prevent a disease or condition in which the activity or expression of PCSK9 has an impact comprising:

(a) administering to a subject a compound that specifically binds PCSK9, preferably an antibody or antigen binding fragment thereof that specifically binds to PCSK9, and

(b) determine which fraction of PCSK9 in the blood is bound to the compound of (a).

Normally, compounds that specifically bind 10% to 100% (preferably 20% to 100%, more preferably 30% to 100%, more preferably 40% to 100%, more preferably 50% to 100%) PCSK9 present in the blood when used in stoichiometric amounts, will be usable to improve, correct, inhibit or prevent a disease or condition in which the activity or expression of PCSK9 has an impact.

Preferably, the disease or condition in which the expression or activity of PCSK9 has an impact is selected from the group consisting of: elevated levels of LDL-C, hypercholesterolemia, hyperlipidemia, dyslipidemia, atherosclerosis and cardiovascular diseases or any of the other diseases described in the first or second aspect.

Antibodies and antigen-binding fragments thereof that can be used to practice the twelfth aspect of the present disclosure are described in the "Preferred antibodies" section.

In a thirteenth aspect, the present disclosure relates to a method of treatment of a disease or condition in which the expression or activity of PCSK9 causes an impact comprising

administering a therapeutic amount of an antibody or an antigen-binding fragment thereof that specifically binds PCSK9h (proprotein convertase subtilisin / human type 9 kexin) to a subject in need thereof,

in which the subject in need thereof is classified into one or more of the following groups of subjects: (i) subjects who have a serum cholesterol level of (C-LDL) of at least 100 mg / dl, preferably at less 130 mg / dl, more preferably at least 160 mg / dl, even more preferably at least 200 mg / dl; (ii) subjects who have a serum C-HDL level below 40 mg / dl; (iii) subjects who have a serum cholesterol level of at least 200 mg / dl, preferably at least 240 mg / dl; (iv) subjects who have a serum triacylglycerols level of at least 150 mg / dl, for example, at least 200 mg / dl or at least 500 mg / dl, in which said triacylglycerols level is determined after fasting during at least 8 hours; (v) subjects who are at least 35 years old, for example, at least 40 years old, at least 45 years old, at least 50 years old, at least 55 years old, at least 60 years old, at least 65 years old, or at least 75 years old; (vi) subjects younger than 75, for example, younger than 70, younger than 65, younger than 60, younger than 55, younger than 50, younger than 45, or younger than 40 years; (vii) subjects who have a BMI of 25 or more (for example, 26 or more, 27 or more, 28 or more, 29 or more, 30 or more, 31 or more, 32 or more, 33 or more, 34 or more, 35 or more, 36 or more, 37 or more, 38 or more, or 39 or more);

(viii) male subjects; (ix) female subjects; (x) subjects in which the administration of said antibody or antigen binding fragment thereof leads to a reduction in serum C-LDL level of at least 30 mg / dl, preferably at least 40 mg / dl, more preferably at least 50 mg / dl, more preferably at least 60 mg / dl, more preferably at least 70 mg / dl, with respect to the predose level; or (xi) subjects in which the administration of said antibody or antigen-binding fragment thereof leads to a reduction in serum C-LDL level of at least 20%, preferably at least 30%, more preferably at least 40%, more preferably at least 50%, more preferably at least 60%, with respect to the predose level.

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In a fourteenth aspect, the present disclosure refers to a method of treatment of a disease or condition in which the expression or activity of PCSK9 causes an impact comprising

administering a therapeutic amount of an antibody or an antigen-binding fragment thereof that specifically binds PCSK9h (proprotein convertase subtilisin / human type 9 kexin) to a subject in need thereof,

in which the subject in need thereof is not classified into one or more of the following groups of subjects: (i) smokers; (ii) persons 70 years of age or older; (iii) people suffering from hypertension; (iv) women who are pregnant; (v) women who are trying to get pregnant; (vi) women who are breastfeeding;

(vii) people who have or have ever had a disease that affects the liver; (viii) people who had some unexplained abnormal blood test for liver function; (ix) people who drink excessive amounts of alcohol; (x) people who have kidney problems; (xi) people suffering from hypothyroidism; (xii) people suffering from muscular disorders; (xiii) people who have had previous muscle problems during treatment with lipid lowering medicine; (xiv) people who have serious problems with their breathing; (xv) people taking one or more of the following medicines: medicines that alter the way the immune systems work (for example, cyclosporine or antihistamines), antibiotics or antifungal medicines (for example, erythromycin, clarithromycin, ketoconazole, itraconazole , rifampicin, fusidic acid), medicines that regulate lipid levels (for example, gemfibrozil, colestipol), calcium channel blockers (for example, verapamil, diltiazem), medicines that regulate heart rhythm (digoxin, amiodarone), protease inhibitors used in the treatment of HIV (for example, nelfinavir), warfarin, oral contraceptives, antacids or St. John's wort; or (xvi) people who drink more than 0.1 l of grapefruit juice per day or who eat more than half a grapefruit per day; (xvii) people who have a body mass index (BMI) greater than 40; (xviii) people who have a body mass index (BMI) of less than 18; (xix) people suffering from type 1 diabetes or type 2 diabetes; (xx) people positive for hepatitis B or hepatitis C; (xxi) people who have a known sensitivity to therapeutic drugs of monoclonal antibodies; (xxii) people who have a neutrophil concentration less than 1500 / mm3; (xxiii) people who have a platelet concentration of less than 100,000 / mm3; (xxiv) males who have a serum creatinine level greater than 1.5 x ULN (upper limit of normal); (xxv) women who have a serum creatinine level greater than 1.4 x ULN (upper limit of normal); (xxvi) people who have a level of alanine transaminase (ALT) or level of aspartate transaminase (AST) greater than 2 x ULN; or (xxvii) people who have a CPK level greater than 3 x ULN.

In preferred embodiments of the thirteenth and fourteenth aspects, the disease or condition in which the expression or activity of PCSK9 causes an impact is improved, corrected, inhibited or prevented with a PCSK9 antagonist.

In preferred embodiments of the thirteenth and fourteenth aspects, the disease or condition in which the expression or activity of PCSK9 causes an impact is selected from the group consisting of: elevated levels of LDL-C, hypercholesterolemia, hyperlipidemia, dyslipidemia, atherosclerosis and cardiovascular diseases or any of the other diseases or conditions described in the other aspects disclosed herein, such as the first or second aspect.

In preferred embodiments of the thirteenth and fourteenth aspect, the subject in need thereof is a subject indicated for apheresis of LDL, a subject with activating mutations of PCSK9, a subject with heterozygous familial hypercholesterolemia, a subject with primary hypercholesterolemia, for example , a subject with primary familial or non-familial primary hypercholesterolemia, a subject with hypercholesterolemia such as primary hypercholesterolemia that is not controlled with statins, a subject at risk of developing hypercholesterolemia, a subject with hypercholesterolemia, a subject with hyperlipidemia, a subject with dyslipidemia, a subject with atherosclerosis or a subject with cardiovascular diseases, or any of the other subjects described in the first or second aspects. Most preferably, the subject in need thereof is a human subject. Additional preferred or suitable subjects are described in the other aspects described herein.

Antibodies and antigen-binding fragments thereof that can be used to implement the thirteenth and fourteenth aspect and the other aspects of the present disclosure are described in the "Preferred antibodies" section.

In preferred embodiments of the thirteenth and fourteenth aspects, the method further comprises: administering a therapeutic amount of an HMG-CoA reductase inhibitor to the subject at a dosage of between 0.05 mg and 100 mg. In some embodiments, the HMG-CoA reductase inhibitor is administered three times per day, twice per day, or once per day. In some embodiments, the HMG-CoA reductase inhibitor is administered every day, every two days, every third day, every fourth day, every fifth day, or every sixth day. In some embodiments, the HMG-CoA reductase inhibitor is administered every week, every two weeks, every three weeks, or every four weeks. In some embodiments, the HMG-CoA reductase inhibitor is administered in the morning, at noon or in the afternoon. In preferred embodiments, the HMG-CoA reductase inhibitor is administered once a day, preferably orally, preferably in the afternoon. Preferably, the HMG-CoA reductase inhibitor is a statin. More preferably, the statin is selected from the group consisting of cerivastatin, atorvastatin, simvastatin, pitavastatin, rosuvastatin, fluvastatin, lovastatin and pravastatin. In realizations

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Additional preferred of the thirteenth and fourteenth aspect, the method comprises administering a therapeutic amount of a statin to the subject, in which the statin is:

- cerivastatin administered at a daily dose of between 0.05 mg and 2 mg, preferably at a daily dose of 0.2 mg, 0.4 mg, or 0.8 mg;

- atorvastatin administered in a daily dose between 2 mg and 100 mg, preferably in a daily dose of 10 mg, 20 mg, 40 mg, or 80 mg;

- simvastatin administered in a daily dose between 2 mg and 100 mg, preferably in a daily dose of 5 mg, 10 mg, 20 mg, 40 mg, or 80 mg;

- pitavastatin administered in a daily dose between 0.2 mg and 100 mg, preferably in a daily dose of 1 mg, 2 mg, 5 mg, 10 mg, or 20 mg;

- rosuvastatin administered in a daily dose between 2 mg and 100 mg, preferably in a daily dose of 5 mg, 10 mg, 20 mg, or 40 mg;

- fluvastatin administered in a daily dose between 2 mg and 100 mg, preferably in a daily dose of 20 mg, 40 mg, or 80 mg;

- lovastatin administered at a daily dose between 2 mg and 100 mg, preferably at a daily dose of 10 mg, 20 mg, 40 mg, or 80 mg; or

- pravastatin administered in a daily dose between 2 mg and 100 mg, preferably in a daily dose of 10 mg, 20 mg, 40 mg, or 80 mg.

A further preferred embodiment of the present disclosure combines the characteristics of the thirteenth aspect and the fourteenth aspect as described herein.

In a fifteenth aspect, the present disclosure relates to an antibody or an antigen-binding fragment thereof that specifically binds PCSK9h (proprotein convertase subtilisin / human type 9 kexin), in which the antibody is characterized by one or more more of the following characteristics after administration to a subject, preferably a human or non-human mammal:

13. Reduction of low density lipoprotein (C-LDL) levels from at least approximately -25% to approximately -40% with respect to a predose level with a sustained reduction for at least a period of 14 days, in wherein the sustained reduction is preferably at least -25% and more preferably at least -30% with respect to a predose level, particularly if administered in a dose of about 40 to about 60 mg, preferably about 45 to about 55 mg and more preferably about 50 mg in a biweekly administration schedule (every two weeks, E2W),

14. Reduction of low density lipoprotein (LDL-C) from at least about -50% to about -65% compared to a predose level with a sustained reduction for at least a period of 14 days, in which the Sustained reduction is preferably at least -40% and more preferably at least -45% with respect to a predose level, particularly if administered in a dose of about 100 mg E2W,

15. Reduction of low density lipoprotein (C-LDL) from at least about -60% to at least about -75% [eg, at least about -60%, at least about -65%, at less about -70 or at least about -75%] with respect to a predose level with a sustained reduction for at least a period of 14 days, in which the sustained reduction is preferably at least -55% and more preferably at least -60% with respect to a predose level, particularly when administered in a dose of approximately 150 mg E2W,

16. Reduction of low density lipoprotein (LDL-C) from at least about 40% to about 75% with respect to a predose level with a sustained reduction for at least a 28-day period, in which the sustained reduction it is preferably at least -35% and more preferably at least -40% with respect to a predose level, particularly when administered in a dose of about 200 mg E4W,

17. Reduction of low density lipoprotein (LDL-C) from at least about -50% to about -75% with respect to a predose level with a sustained reduction for at least a 28-day period, in which the Sustained reduction is preferably at least -40% and more preferably at least -45% with respect to a predose level, particularly when administered in a dose of approximately 300 mg E4W,

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18. Increased serum HDL cholesterol levels of at least 2%, at least 2.5%, at least 3%, at least 3.5%, at least 4%, at least 4.5%, at least 5% or at least 5.5% with respect to a predose level, particularly when administered in a dose of approximately 150 mg E2W,

19. reduction of total serum cholesterol from at least about 25% to about 35% with respect to a predose level with a sustained reduction for at least a 24-day period,

20. reduction of total serum cholesterol from at least about 65% to about 80% with respect to a predose level with a sustained reduction for at least a 24-day period,

21. reduction of serum triglyceride levels from at least about 25% to about 40% with respect to a predose level,

22. little or no measurable effect on liver function, as determined by measurements of ALT and AST,

23. little or no measurable effect on troponin levels,

24. increase in one or more of: total cholesterol levels, ApoB levels, C-non HDL levels, Apo-B / ApoA-1 ratio,

The fifteenth aspect antibody of the present disclosure exhibits the above properties preferably if administered in combination with an HMG-CoA reductase inhibitor treatment. Preferred embodiments of the HMG-CoA reductase inhibitors to be used in conjunction with the antibody disclosed herein and dosage and administration guidelines thereof can be found throughout the specification, particularly as described in aspects 1, 2 or 19.

According to a preferred embodiment of the antibodies and antigen binding fragments thereof of the present disclosure, particularly of the antibody or antigen binding fragment according to the fifteenth aspect, the antibody or antigen binding fragment thereof has one or more of the following characteristics:

(i) The antibody or antigen binding fragment comprises heavy and light chain CDRs of a pair of HCVR / LCVR amino acid sequences as shown in SEQ ID NO: 90/92.

(ii) The antibody or antigen binding fragment thereof comprises a pair of HCVR / LCVR amino acid sequences as shown in SEQ ID NO: 90/92.

(iii) The antibody or antigen binding fragment thereof competes to bind PCSK9h with an antibody or antigen binding fragment comprising a pair of amino acid sequences of HCVR / LCVR as shown in SEQ ID NO: 90/92 .

According to another preferred embodiment of the antibodies and antigen binding fragments thereof of the present disclosure, particularly of the antibody or antigen binding fragment according to the fifteenth aspect, the antibody or antigen binding fragment thereof has one or more of the following characteristics:

(i) overcome statin resistance in mammals, especially in rodents such as hamster

(ii) increased expression of LDLR in mammals, particularly in rodents such as hamsters

(iii) decrease in serum LDL-C in rodents such as hamsters

(iv) synergistic decrease of C-LDL in conjunction with the administration of HMG-CoA reductase inhibitor, particularly in rodents such as hamster, in which the HMG-CoA reductase inhibitor is preferably atorvastatin.

Additional suitable structural features and characteristics of the antibody of the present disclosure and particularly of the fifteenth aspect antibody, in addition to antibodies and antigen binding fragments thereof that can be used to practice the fifteenth aspect and the other aspects disclosed in This document is described in the "Preferred antibodies" section.

The antibody of the present disclosure, such as the antibody according to the fifteenth aspect, is preferably formulated as a pharmaceutically administrable formulation as is known in the art, and specifically as described herein, such as dry formulation for solution or formulation. liquid, for example, as described in the twenty-first or twenty-second aspect.

In a sixteenth aspect, the present disclosure relates to an antibody or an antigen-binding fragment thereof that specifically binds PCSK9h (proprotein convertase subtilisin / human type 9 kexin) for use in the treatment of a disease or condition in which the expression or activity of PCSK9 causes an impact,

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wherein the antibody or antigen binding fragment thereof is for administration in a dose of about 50 to 500 mg.

Preferred embodiments of the sixteenth aspect of the present disclosure are described in the fifteenth aspect. Antibodies and antigen-binding fragments thereof that can be used to practice the sixteenth aspect disclosed herein are described in the "Preferred antibodies" section.

According to another preferred embodiment of the sixteenth aspect, the antibody or antigen binding fragment thereof is for administration in a dose of approximately 50, 100, 150, 200, 250, 300, 350, 400, 450 or 500 mg and preferably of approximately 150, 200 or 300 mg.

According to another preferred embodiment of the sixteenth aspect, the disease or condition is selected from the group consisting of: elevated levels of total cholesterol, elevated levels of low density lipoprotein (LDL-C), hypercholesterolemia, hyperlipidemia, dyslipidemia and atherosclerosis, particularly hypercholesterolemia primary, familial hypercholesterolemia or hypercholesterolemia that is not controlled by statins.

According to another preferred embodiment of the sixteenth aspect, the antibody or antigen binding fragment thereof is administered to the subject every two weeks (E2W), every four weeks (E4W) or once a month.

According to another preferred embodiment of the sixteenth aspect, the antibody or antigen binding fragment thereof has one or more of the following characteristics:

(i) It is for use in reducing the levels of low density lipoprotein (LDL-C) from at least approximately -25% to approximately -40% with respect to a predose level with a sustained reduction for at least a period of 14 days, in which the sustained reduction is preferably at least -25% and more preferably at least -30% with respect to a predose level, in which the antibody or antigen-binding fragment thereof is preferably administered in a dose of about 40 to about 60 mg, about 45 to about 55 mg or about 50 mg E2W.

(ii) It is for use in the reduction of low density lipoprotein (LDL-C) from at least about -50% to about -65% with respect to a predose level with a sustained reduction for at least a period of 14 days, in which the sustained reduction is preferably at least -40% and more preferably at least -45% with respect to a predose level, in which the antibody or fragment thereof is preferably administered in a dose of approximately 100 mg E2W.

(iii) It is for use in the reduction of low density lipoprotein (C-LDL) from at least about -60% to at least about -75% [eg, at least about -60%, at least approximately -65%, at least approximately -70 or at least approximately -75%] with respect to a predose level with a sustained reduction for at least a period of 14 days, in which the sustained reduction is preferably at least -55% and more preferably at least -60% with respect to a predose level, in which the antibody or fragment thereof is preferably administered in a dose of approximately 150 mg E2W.

(iv) It is for use in the reduction of low density lipoprotein (LDL-C) from at least about 40% to about 75% with respect to a predose level with a sustained reduction for at least a period of 28 days , wherein the sustained reduction is preferably at least -35% and more preferably at least -40% with respect to a predose level, in which the antibody or fragment thereof is preferably administered in a dose of approximately 200 mg E4W

(v) It is for use in the reduction of low density lipoprotein (LDL-C) from at least about -50% to about -75% with respect to a predose level with a sustained reduction for at least a period of 28 days, in which the sustained reduction is preferably at least -40% and more preferably at least -45% with respect to a predose level, in which the antibody or fragment thereof is preferably administered in a dose of approximately 300 mg E4W.

(vi) It is for use in increasing serum HDL cholesterol levels of at least 2%, at least the

2.5%, at least 3%, at least 3.5%, at least 4%, at least 4.5%, at least 5% or at least

5.5% with respect to a predose level.

(vii) It is for use in reducing total serum cholesterol at least about 25% to about 35% with respect to a predose level with sustained reduction for at least a 24-day period.

(viii) It is for use in the reduction of total serum cholesterol at least about 65% to about 80% with respect to a predose level with sustained reduction for at least a 24-day period.

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25. It is for use in reducing serum triglyceride levels from at least about 25% to about 40% with respect to a predose level.

26. It has little or no measurable effect on liver function, as determined by measurements of ALT and AST, or on troponin levels.

27. It is for use in increasing one or more of: total cholesterol levels, ApoB levels, C-non HDL levels, Apo-B / ApoA-1 ratio.

According to another preferred embodiment of the sixteenth aspect, the antibody or antigen binding fragment thereof is for use in conjunction with an HMG-CoA reductase inhibitor, wherein the HMG-CoA reductase inhibitor is preferably administered in an amount of dose in the range of about 0.05 mg to about 100 mg and is preferably a statin, in which the statin is preferably selected from the group consisting of: cerivastatin, atorvastatin, simvastatin, pitavastatin, rosuvastatin, fluvastatin, lovastatin or pravastatin.

According to another preferred embodiment of the sixteenth aspect, the statin is administered according to one or more of the following dosage or administration guidelines:

(i) statin is administered once a day,

(ii) the statin is administered at a dose of about 0.5 to about 100 mg, about 5 to about 90 mg, of about 10, 20, 40 or 80 mg and is preferably atorvastatin.

Additional dosage and administration guidelines for the antibody, antigen fragment thereof or HMG-CoA reductase inhibitor are described in the other aspects disclosed herein and preferably in the first, second or tenth ninth aspect.

In a seventeenth aspect, the present disclosure relates to an antibody or antigen-binding fragment thereof that specifically binds PCSK9h (proprotein convertase subtilisin / human type 9 kexin) for use in the treatment of a disease or condition in which the expression or activity of PCSK9 causes an impact,

wherein the antibody or antigen binding fragment thereof is for administration to a subject that is in at least one of the following groups of subjects: (i) subjects having a cholesterol level of (C-LDL) in serum of at least 100mg / dl, preferably at least 130 mg / dl, more preferably at least 160 mg / dl, even more preferably at least 200 mg / dl; (ii) subjects who have a serum HDL-C level below 40 mg / dl; (iii) subjects who have a serum cholesterol level of at least 200 mg / dl, preferably at least 240 mg / dl; (iv) subjects who have a serum triacylglycerols level of at least 150 mg / dl, for example, at least 200 mg / dl or at least 500 mg / dl, in which said triacylglycerols level is determined after fasting during at least 8 hours; (v) subjects who are at least 35 years old, for example, at least 40 years old, at least 45 years old, at least 50 years old, at least 55 years old, at least 60 years old, at least 65 years old, or at least 75 years old; (vi) subjects younger than 75, for example, younger than 70, younger than 65, younger than 60, younger than 55, younger than 50, younger than 45, or younger than 40 years; (vii) subjects who have a BMI of 25 or more (for example, 26 or more, 27 or more, 28 or more, 29 or more, 30 or more, 31 or more, 32 or more, 33 or more, 34 or more, 35 or more, 36 or more, 37 or more, 38 or more, or 39 or more); (viii) male subjects; (ix) female subjects; (x) subjects in which the administration of said antibody or antigen binding fragment thereof leads to a reduction in serum C-LDL level of at least 30 mg / dl, preferably at least 40 mg / dl, more preferably at least 50 mg / dl, more preferably at least 60 mg / dl, more preferably at least 70 mg / dl, with respect to the predose level; or (xi) subjects in which the administration of said antibody or antigen-binding fragment thereof leads to a reduction in serum C-LDL level of at least 20%, preferably at least 30%, more preferably at least 40%, more preferably at least 50%, more preferably at least 60%, with respect to the predose level.

In an eighteenth aspect, the present disclosure relates to an antibody or antigen binding fragment thereof that specifically binds PCSK9h (proprotein convertase subtilisin / human type 9 kexin) for use in the treatment of a disease or condition in which the expression or activity of PCSK9 causes an impact,

wherein the antibody or antigen binding fragment thereof is for administration to a subject that is not classified into one or more of the following groups of subjects: (i) smokers; (ii) persons 70 years of age or older; (iii) people suffering from hypertension; (iv) women who are pregnant; (v) women who are trying to get pregnant; (vi) women who are breastfeeding; (vii) people who have or have ever had a disease that affects the liver; (viii) people who had some unexplained abnormal blood test for liver function; (ix) people who drink excessive amounts of alcohol; (x) people who have kidney problems; (xi) people suffering from hypothyroidism; (xii) people suffering from muscular disorders; (xiii) people who have had previous muscle problems during treatment with lipid lowering medicine; (xiv) people who have serious problems with their breathing; (xv) people taking one or more of the following medicines:

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medicines that alter the way immune systems work (for example, cyclosporine or antihistamines), antibiotics or antifungal medicines (for example, erythromycin, clarithromycin, ketoconazole, itraconazole, rifampin, fusidic acid), medicines that regulate lipid levels (for example, gemfibrozil, colestipol), calcium channel blockers (for example, verapamil, diltiazem), medicines that regulate heart rate (digoxin, amiodarone), protease inhibitors used in the treatment of HIV (for example, nelfinavir), warfarin, oral contraceptives, antacids or St. John's wort; or (xvi) people who drink more than 0.1 l of grapefruit juice per day or who eat more than half a grapefruit per day; (xvii) people who have a body mass index (BMI) greater than 40; (xviii) people who have a body mass index (BMI) of less than 18; (xix) people suffering from type 1 diabetes or type 2 diabetes; (xx) people positive for hepatitis B or hepatitis C; (xxi) people who have a known sensitivity to therapeutic drugs of monoclonal antibodies; (xxii) people who have a neutrophil concentration less than 1500 / mm3; (xxiii) people who have a platelet concentration of less than 100,000 / mm3; (xxiv) males who have a serum creatinine level greater than 1.5 x ULN (upper limit of normal); (xxv) women who have a serum creatinine level greater than 1.4 x ULN (upper limit of normal); (xxvi) people who have a level of alanine transaminase (ALT) or level of aspartate transaminase (AST) greater than 2 x ULN; or (xxvii) people who have a CPK level greater than 3 x ULN.

In preferred embodiments of the fifteenth to eighteenth aspect, the disease or condition in which the expression or activity of PCSK9 causes an impact is improved, corrected, inhibited or prevented with a PCSK9 antagonist.

In preferred embodiments of the fifteenth to eighteenth aspect, the disease or condition in which the expression or activity of PCSK9 causes an impact is selected from the group consisting of: elevated levels of LDL-C, hypercholesterolemia, hyperlipidemia, dyslipidemia, atherosclerosis and cardiovascular diseases, or any other of the diseases and conditions described in the first or second aspect.

In preferred embodiments of the fifteenth to eighteenth aspect, the antibody or antigen binding fragment thereof is for administration to a subject indicated for LDL apheresis, a subject with PCSK9 activating mutations, a subject with heterozygous familial hypercholesterolemia, a subject with primary hypercholesterolemia, for example, a subject with primary or non-familial primary family hypercholesterolemia, a subject with hypercholesterolemia such as primary hypercholesterolemia that is not controlled with statins, a subject at risk of developing hypercholesterolemia, a subject with hypercholesterolemia, a subject with hyperlipidemia , a subject with dyslipidemia, a subject with atherosclerosis or a subject with cardiovascular disease or any other subject as described in the first or second aspect. Most preferably, the subject is a human subject.

Antibodies and antigen-binding fragments thereof that can be used to practice the fifteenth to eighteenth aspect of the present disclosure are described in the "Preferred antibodies" section.

In preferred embodiments of the 18th to 18th aspect, the antibody or antigen binding fragment thereof is for administration in combination with a dosage of between 0.05 mg and 100 mg of an HMG-CoA reductase inhibitor. In some embodiments, the HMG-CoA reductase inhibitor will be administered three times a day, twice a day, or once a day. In some embodiments, the HMG-CoA reductase inhibitor will be administered every day, every two days, every third day, every fourth day, every fifth day, or every sixth day. In some embodiments, the HMG-CoA reductase inhibitor will be administered every week, every two weeks, every three weeks, or every four weeks. In some embodiments, the HMG-CoA reductase inhibitor will be administered in the morning, at noon or in the afternoon. In preferred embodiments, the HMG-CoA reductase inhibitor is to be administered once a day, preferably orally, preferably in the afternoon. Preferably, the HMG-CoA reductase inhibitor is a statin. More preferably, the statin is selected from the group consisting of cerivastatin, atorvastatin, simvastatin, pitavastatin, rosuvastatin, fluvastatin, lovastatin and pravastatin. In further preferred embodiments of the 18th to 18th aspect, the antibody or antigen binding fragment thereof is for administration in combination with a statin, wherein the statin is

- cerivastatin to be administered at a daily dose of between 0.05 mg and 2 mg, preferably at a daily dose of 0.2 mg, 0.4 mg, or 0.8 mg;

- atorvastatin to be administered in a daily dose between 2 mg and 100 mg, preferably in a daily dose of 10 mg, 20 mg, 40 mg, or 80 mg;

- simvastatin to be administered in a daily dose between 2 mg and 100 mg, preferably in a daily dose of 5 mg, 10 mg, 20 mg, 40 mg, or 80 mg;

- pitavastatin to be administered in a daily dose between 0.2 mg and 100 mg, preferably in a daily dose of 1 mg, 2 mg, 5 mg, 10 mg, or 20 mg;

- rosuvastatin to be administered at a daily dose between 2 mg and 100 mg, preferably at a daily dose of 5 mg, 10 mg, 20 mg, or 40 mg;

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- fluvastatin to be administered in a daily dose between 2mg and 100 mg, preferably in a daily dose of 20 mg, 40 mg, or 80 mg;

- lovastatin to be administered in a daily dose between 2 mg and 100 mg, preferably in a daily dose of 10 mg, 20 mg, 40 mg, or 80 mg; or

- pravastatin to be administered in a daily dose between 2 mg and 100 mg, preferably in a daily dose of 10 mg, 20 mg, 40 mg, or 80 mg.

A further preferred embodiment of the present disclosure combines the characteristics of the sixteenth and seventeenth or eighteenth and eighteenth or seventeenth and eighteenth or eighteenth and seventeenth and eighteenth aspects as described herein.

In a nineteenth aspect, the present disclosure relates to a pharmaceutical composition comprising the antibody or antigen-binding fragment thereof disclosed herein together with an excipient.

0 pharmaceutically acceptable vehicle.

According to a preferred embodiment of the tenth ninth aspect, the antibody or fragment thereof is as described in the fifteenth, first or second aspect; Additional suitable characteristics of the antibody are described in the "Preferred antibodies" section.

According to another preferred embodiment, the pharmaceutical composition comprises about 40 to about 500 mg of the antibody or antigen binding fragment per dose.

According to another preferred embodiment, the pharmaceutical composition comprises about 50 mg to about 500 mg, about 50 mg to about 300 mg, about 50 mg, about 100 mg, about 150 mg, about 200 mg, about 250 mg, about 300 mg, approximately 350 mg, approximately 400 mg, approximately 450 mg or approximately 500 mg of the antibody or antigen binding fragment thereof.

According to another preferred embodiment, the pharmaceutical composition comprises about 150, 200 or 300 mg of the antibody or antigen binding fragment thereof.

According to another preferred embodiment, the pharmaceutical composition comprises an effective dose of an antibody or an antigen-binding fragment thereof that specifically binds PCSK9h (proprotein convertase subtilisin / human type 9 kexin), wherein the dose is sufficient for reduction. Sustained low density lipoprotein (C-LDL) levels for a period of at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at at least 22, at least 23 or at least 28 days after administration, together with a pharmaceutically acceptable carrier or excipient. According to another preferred embodiment, the dose is sufficient for sustained reduction of LDL-C levels for a period of at least 14 days, 28 days or

1 month.

According to another preferred embodiment, the pharmaceutical composition comprises an effective amount of an HMG-CoA reductase inhibitor.

According to another preferred embodiment, the pharmaceutical composition is disposed together with an effective amount of an HMG-CoA reductase inhibitor.

According to another preferred embodiment, the HMG-CoA reductase inhibitor is a statin, preferably selected from the list consisting of: cerivastatin, atorvastatin, simvastatin, pitavastatin, rosuvastatin, fluvastatin, lovastatin or pravastatin and is preferably atorvastatin.

According to another preferred embodiment, the pharmaceutical composition comprises about 0.05 mg to about 100 mg, about 0.5 mg to about 100 mg, about 5 mg to about 90 mg, about 10 mg, about 20 mg, about 40 mg or about 80 mg of HMG-CoA reductase inhibitor and preferably about 10, about 20, about 40 or about 80 mg.

According to another preferred embodiment, the pharmaceutical composition comprises an effective dose of HMG-CoA reductase inhibitor to reduce LDL-D levels by administration once a day.

The antibody or antigen-binding fragment thereof to be used for the pharmaceutical composition according to the present disclosure may be any antibody as described herein, such as in the "Preferred antibodies" or in the fifteenth and other aspects. additional.

According to a preferred embodiment, the antibody or antigen binding fragment thereof has one or more of the following characteristics when administered to a subject, such as a human or non-human mammal:

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to. reduction of low density lipoprotein (C-LDL) levels from at least about -25% to about -40% compared to a predose level with a sustained reduction for at least a period of 14 days after administration to a subject, in which the sustained reduction is preferably at least -25% and more preferably at least -30% with respect to a predose level, particularly if administered in a dose of about 40 to about 60 mg, preferably about 45 to about 55 mg and more preferably about 50 mg in a biweekly administration schedule (every two weeks, E2W);

b. reduction of low density lipoprotein (LDL-C) from at least about -50% to about -65% with respect to a predose level with a sustained reduction for at least a period of 14 days after administration to a subject, wherein the sustained reduction is preferably at least -40% and more preferably at least -45% with respect to a predose level, particularly if administered in a dose of about 100 mg E2W.

C. reduction of low density lipoprotein (C-LDL) from at least about -60% to at least about -75% [eg, at least about -60%, at least about -65%, at least about -70 or at least about -75%] with respect to a predose level with a sustained reduction for at least a period of 14 days after administration to a subject, in which the sustained reduction is preferably at least -55 % and more preferably at least -60% with respect to a predose level, particularly when administered in a dose of approximately 150 mg E2W,

d. reduction of low density lipoprotein (C-LDL) of at least about 40% at

approximately 75% with respect to a predose level with a sustained reduction for at least a period of 28 days, in which the sustained reduction is preferably at least -35% and more preferably at least -40% with respect to a predose level, particularly when administered at a dose of approximately 200 mg E4W

and. reduction of low density lipoprotein (C-LDL) of at least about -50% at

approximately -75% with respect to a predose level with a sustained reduction for at least a period of 28 days after administration to a subject, in which the sustained reduction is preferably at least -40% and more preferably at least -45% compared to a predose level, particularly when administered in a dose of approximately 300 mg E4W,

F. increased serum HDL cholesterol levels of at least 2%, at least 2.5%, at least

3%, at least 3.5%, at least 4%, at least 4.5%, at least 5% or at least 5.5% with respect

at a predose level after administration to a subject, particularly when administered in a dose of approximately 150 mg E2W,

g. little or no measurable effect on troponin levels after administration to a subject,

h. increase in one or more of: total cholesterol levels, ApoB levels, C-non HDL levels, Apo-B / ApoA-1 ratio, after administration to a subject.

According to another preferred embodiment, the antibody or antigen binding fragment thereof is capable of overcoming statin resistance when administered to a subject with statin resistant hypercholesterolemia.

According to another preferred embodiment, the antibody or antigen binding fragment thereof comprises the heavy and light chain CDRs of a pair of HCVR / LCVR amino acid sequences as shown in SEQ ID NO: 90/92 or a pair of substantially identical sequences that have at least 98% or 99% identity with them.

According to another preferred embodiment, the antibody or antigen binding fragment thereof comprises a pair of HCVR / LCVR amino acid sequences as shown in SEQ ID NO: 90/92 or a pair of substantially identical sequences having at least 98 % or 99% identity with them.

According to another preferred embodiment, the antibody or antigen binding fragment thereof competes to bind PCSK9h with an antibody or antigen binding fragment comprising a pair of amino acid sequences of HCVR / LCVR as shown in SEQ ID NO: 90 / 92.

According to another preferred embodiment, the antibody or antigen binding fragment thereof binds an epitope comprising amino acid residue 238 of PCSK9h (SEQ ID NO: 755).

According to another preferred embodiment, the antibody or antigen binding fragment thereof binds an epitope comprising one or more of the amino acid residues at positions 238, 153, 159 and 343 of PCSK9h (SEQ ID NO: 755).

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According to another preferred embodiment, the antibody or antigen binding fragment thereof binds an epitope that does not comprise an amino acid residue at positions 192, 194, 197 and / or 237 of PCSK9h (SEQ ID NO: 755).

The pharmaceutical composition may be formulated according to any pharmaceutically administrable formulation as is known in the art, and specifically as described herein, such as dry formulation for solution or liquid formulation. Suitable antibody formulations are known in the art and comprise dry formulations (e.g., freeze dried, spray dried or lyophilized, concentrated without water), in addition to liquid formulations (e.g., solutions). Suitable statin formulations are well known in the art and comprise dry formulations, in addition to liquid formulations, for example, suspensions, dispersions and solutions (for a reference see, for example, "Statins therapy: a review on conventional and novel formulation approaches" R. Tiwari and K. Pathak, Journal of Pharmacy and Pharmacology, 2011, which is incorporated herein in its entirety).

According to a preferred embodiment, the pharmaceutical composition comprises the antibody or antigen-binding fragment thereof as a dry formulation for dissolution such as a lyophilized powder, freeze-dried or spray-dried powder or concentrate without water.

According to another preferred embodiment, the pharmaceutical composition comprises the antibody or antigen binding fragment thereof as a liquid formulation, for example, solution for injection or infusion.

According to another preferred embodiment, the pharmaceutical composition comprises the HMG-CoA reductase inhibitor as an oral or peroral formulation, for example, capsule or tablet, or as a liquid formulation, for example, suspension, dispersion or solution, for example, for peroral administration, injection or infusion

According to another preferred embodiment, the pharmaceutical composition is for use in the treatment of a disease or disorder for use in the treatment of a disease or condition in which the expression or activity of PCSK9 causes an impact or to reduce elevated levels of Total or elevated C-LDL cholesterol. Further preferred uses, dosage regimens, administration guidelines for the antibody or fragment thereof or for the HMG-CoA reductase inhibitor, or populations to be treated with the pharmaceutical composition are described in the present application, for example, in the other aspects disclosed. in this document such as the first or second aspect.

In a twentieth aspect, the present disclosure relates to a solution for injection as described herein comprising the antibody or antigen binding fragment thereof disclosed herein, and preferably comprising about 40 mg to about 200 mg or about 50 to about 200 mg, for example, about 40 mg, about 50 mg, about 75 mg, about 100 mg, about 150 mg or about 200 mg of the antibody or antigen binding fragment thereof per 1 ml volume .

In a twenty-first aspect, the present disclosure relates to a dry formulation as described herein comprising the antibody or antigen binding fragment thereof disclosed herein, and preferably comprising about 40 mg to about 500 mg, 50 to about 500 mg, about 50 to about 400, about 50 to about 300 for example, about 40 mg, about 50 mg, about 75 mg, about 100 mg, about 150 mg, about 200 mg, about 250 mg, approximately 300 mg, approximately 350 mg, approximately 400 mg, approximately 450 mg or approximately 500 mg and more preferably approximately 50 mg, approximately 100 mg, approximately 150 mg, approximately 200 mg, approximately 250 mg, approximately 300 mg and even more preferably approximately 150 mg, approximately 200 mg or approximately 300 mg of the antibody or antigen binding fragment thereof by dose.

The formulations of the present disclosure may comprise additional active ingredients such as an HMG-CoA reductase inhibitor as described herein. Preferred embodiments of the 20th or 21st aspect are described in other sections of the present application, for example, in the other aspects disclosed herein such as the fifteenth, tenth, ninth or twentieth aspects.

Suitable antibody formulations are generally known in the art and comprise dry formulations (e.g., freeze dried, spray dried or lyophilized, concentrated without water), in addition to liquid formulations (e.g., solutions). Suitable statin formulations are well known in the art and comprise dry formulations, in addition to liquid formulations, for example, suspensions, dispersions and solutions (for a reference see, for example, "Statins therapy: a review on conventional and novel formulation approaches" R. Tiwari and K. Pathak, Journal of Pharmacy and Pharmacology, 2011, which is incorporated herein in its entirety).

According to a twenty-second aspect, the present disclosure relates to an antibody or antigen-binding fragment thereof as comprised in one of the pharmaceutical compositions according to the tenth ninth aspect.

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In a twenty-third aspect, the present disclosure relates to a unit dosage form comprising the antibody, antigen-binding fragment thereof or pharmaceutical composition disclosed herein. Suitable embodiments of the antibody, pharmaceutical composition or formulation to be used to implement the twenty-third aspect of the present disclosure can be obtained from the respective sections of the present application, such as the first, second, tenth, ninth, twenty-first, twenty-first or twenty-second aspects of the "Preferred antibodies" section.

According to a preferred embodiment, the unit dosage form comprises approximately 40 mg, approximately 50 mg, approximately 75 mg, at approximately 100 mg, approximately 150 mg, approximately 200 mg, approximately 250 mg, approximately 300 mg, approximately 350 mg, approximately 400 mg, about 450 mg, or about 500 mg of the antibody or antigen binding fragment thereof.

According to another preferred embodiment, the unit dosage form comprises the antibody or fragment thereof as a dry formulation for dissolution in a hermetically sealed container such as a vial, a vial or envelope.

According to another preferred embodiment, the unit dosage form comprises the antibody or fragment thereof as a liquid formulation in a hermetically sealed container such as a vial, an envelope, a pre-filled syringe, pre-filled auto-injector or a cartridge for a reusable syringe or applicator.

According to another preferred embodiment, the amount of active ingredient is indicated in the hermetically sealed container.

As used in the different aspects and embodiments of the present disclosure and particularly in the twenty-third aspect, the term "unit dosage form" refers to physically discrete units suitable as unit dosages for human and animal subjects, each unit containing a quantity predetermined active material (for example, about 40 mg or about 50 mg to about 500 mg of antibody against PCSK5 and / or of, for example, 0.05 mg to 100 mg of HMG-CoA reductase inhibitor) calculated to produce the desired therapeutic effect in association with the diluent, carrier or pharmaceutical carrier required. The specifications for the novel unit dosage forms of the present disclosure are imposed by and are directly dependent on (a) the unique characteristics of the active material and the particular therapeutic effect to be achieved, and (b) the limitation inherent in the subject of combining an active material such for therapeutic use in animals or humans, as disclosed in this specification, these being characteristics of the present disclosure. Examples of suitable unit dosage forms according to the present disclosure are vials, tablets, capsules, troches, suppositories, powder containers, wafers, seals, ampoules, segregated multiples of any of the foregoing, and other forms as described herein. or generally known in the art.

One or more such unit dosage forms of the antibody may be comprised in an article of manufacture of the present disclosure, which optionally further comprises one or more unit dosage forms of an HMG-CoA reductase inhibitor (eg, a honeycomb container of tablets comprising as active ingredient the HMG-CoA reductase inhibitor).

The term "active material" refers to any material with therapeutic activity, such as one or more active ingredients. The active ingredients to be used as therapeutic agents can be easily prepared in such a unit dosage form with the use of pharmaceutical materials that are themselves available in the art and can be prepared by established procedures. Preferred active ingredients of the present disclosure are the antibody or fragment thereof or an HMG-CoA reductase inhibitor such as a statin.

In a preferred embodiment, the unit dosage form comprises 40 - about 500 mg of the antibody or an antigen binding fragment of the present disclosure. According to another preferred embodiment, the unit dosage form comprises approximately 40 mg, approximately 50 mg, approximately 75 mg, approximately 100 mg, approximately 150 mg, approximately 200 mg, approximately 250 mg, approximately 300 mg, approximately 350 mg, approximately 400 mg , about 450 mg, or about 500 mg and more preferably about 150 mg, about 200 mg or about 300 mg of the antibody or antigen binding fragment thereof. Additional preferred dosages, and dosage guidelines, are as described anywhere in the application, such as in the first, second or fifteenth to nineteenth aspects.

According to another preferred aspect, the unit dosage form comprises the antibody, antigen-binding fragment thereof or pharmaceutical composition as a dry formulation for solution such as a lyophilized powder, freeze dried powder or concentrated without water. According to another preferred embodiment, the dried formulation is comprised in a hermetically sealed container such as a vial, a vial or envelope.

According to another preferred embodiment, the unit dosage form comprises the antibody, antigen binding fragment thereof or pharmaceutical composition as a liquid formulation, for example, solution for injection or infusion. According to another preferred embodiment, the liquid formulation is comprised in a hermetically sealed container such as a vial, an envelope, a pre-filled syringe, a pre-filled auto-injector or a cartridge for a reusable syringe or applicator.

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It is further preferred, if the amount of active ingredient (eg antibody) is indicated on the hermetically sealed package of the unit dosage form.

The following preparations are illustrative of the preparation of the unit dosage forms of the present disclosure, and not as a limitation thereof. Various dosage forms can be prepared that are integrated into the present disclosure. For example, a unit dosage per vial may contain 0.5 ml, 1 ml, 2 ml, 3 ml, 4 ml, 5 ml, 6 ml, 7 ml, 8 ml, 9 ml, 10 ml, 15 ml, or 20 ml of antibody against PCSK5 or a fragment thereof ranging from about 40 to about 500 mg of antibody against PCSK5. If necessary, these preparations can be adjusted to a desired concentration by adding a sterile diluent to each vial.

In one embodiment, the components of the disclosure formulation are supplied either separately or mixed together in unit dosage form, for example, as a dry solution formulation or a liquid formulation. The preparation of pharmaceutically acceptable formulations of proteinaceous biomolecules such as antibodies or antigen binding fragments thereof or of small molecule compounds such as statins is generally known in the art. According to a preferred embodiment, the active ingredients, active material or pharmaceutical composition according to the present disclosure is a dry formulation for liquid dissolution, such as a lyophilized powder, freeze dried powder or concentrated without water, preferably comprised in a hermetically sealed container such as a vial, a vial or envelope, and preferably indicating the amount of active agent. Where the composition is to be administered by infusion, it can be dispensed with an infusion bottle containing sterile pharmaceutical grade water or saline solution. Where the composition is administered by injection, a sterile water vial for injection or saline solution can be provided so that the components can be mixed before administration.

Disclosure formulations include bulk drug compositions useful in the manufacture of pharmaceutical compositions (eg, compositions that are suitable for administration to a subject or patient) that can be used in the preparation of unit dosage forms. In a preferred embodiment, a composition of the disclosure is a pharmaceutical composition. Such compositions comprise a prophylactically or therapeutically effective amount of one or more prophylactic or therapeutic agents (for example, an antibody disclosed herein or another prophylactic or therapeutic agent), and a pharmaceutically acceptable carrier. Preferably, the pharmaceutical compositions are formulated to be suitable for the route of administration to a subject.

In a specific embodiment, the term "pharmaceutically acceptable" means authorized by a US federal government regulatory agency. or state or AEM (European Medicines Agency) or listed in the US Pharmacopoeia. (United States Pharmacopeia-33 / National Formulary-28 Reissue, published by United States Pharmacopeia Convention, Inc., Rockville Md., Publication date: April 2010) or other pharmacopoeia generally recognized for use in animals, and more particularly in Humans.

The term "vehicle" refers to a diluent, adjuvant {for example, Freund's adjuvant (complete and incomplete)), excipient, or vehicle with which the therapeutic drug is administered. Such pharmaceutical vehicles may be sterile liquids, such as water and oils, which include those of petroleum, animal, vegetable or synthetic origin, such as peanut oil, soybean oil, mineral oil, sesame oil. Water is a preferred vehicle when the pharmaceutical composition is administered intravenously. Saline solutions and aqueous solutions of dextrose and glycerol can also be used as liquid carriers, particularly for injectable solutions. Suitable pharmaceutical excipients include starch, glucose, lactose, sucrose, gelatin, malt, rice, flour, limestone, silica gel, sodium stearate, glycerol monostearate, talc, sodium chloride, skimmed milk powder, glycerol, propylene, glycol, water, ethanol The composition, if desired, may also contain minor amounts of wetting or emulsifying agents, or pH buffering agents. These compositions can take the form of solutions, suspensions, emulsion, tablets, pills, capsules, powders, sustained release formulations. The oral formulation may include standard vehicles such as pharmaceutical grades of mannitol, lactose, starch, magnesium stearate, sodium saccharin, cellulose, magnesium carbonate, etc. Examples of suitable pharmaceutical vehicles are described in "Remington's Pharmaceutical Sciences" by E. W. Martin. Such compositions will contain a prophylactically or therapeutically effective amount of the antibody, preferably in purified form, together with a suitable amount of vehicle so as to provide the form for proper administration to the patient. The composition may also contain one or more other active ingredients such as an HMG-CoA reductase inhibitor. The formulation must be adapted to the mode of administration.

Generally, the components of compositions of the disclosure are supplied either separately or mixed together in unit dosage form, for example, as a dry formulation for dissolution such as a lyophilized powder, freeze dried powder or concentrated without water in a container Hermetically sealed such as a blister or envelope that indicates the amount of active agent. The components of the disclosed compositions may also be supplied as a mixed liquid formulation (i.e. solution for injection or infusion) in a hermetically sealed container such as a vial, envelope, a pre-filled syringe or auto-injector, or a cartridge for a syringe or applicator reusable (for example, pen or auto-injector). Where the composition is to be administered by infusion, it can be dispensed with an infusion bottle containing water.

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Sterile pharmaceutical grade or saline solution. Where the composition is administered by injection, a sterile water vial for injection or saline solution can be provided so that the components can be mixed before administration. The composition may also comprise two or more active ingredients that are each formulated in a different way or in the same manner, for example, a combination of an antibody disclosed herein together with an HMG-CoA reductase inhibitor disclosed in the present document

The disclosure also provides that the formulation is packaged in a hermetically sealed container such as a blister or envelope indicating the amount of antibody. In one embodiment, the formulation disclosed herein comprising an antibody is supplied as a dry sterilized lyophilized powder, freeze dried powder or concentrated without water in a hermetically sealed container and can be reconstituted, for example, with water or saline solution. the appropriate concentration for administration to a subject. In one embodiment, the formulation disclosed herein comprising an antibody is supplied as a dry sterile lyophilized powder in a tightly sealed container at a unit dosage of at least 40 mg, at least 50 mg, more preferably at least 75 mg, at least 100 mg, at least 150 mg, at least 200 mg, at least 250 mg, at least 300 mg, at least 350 mg, at least 400 mg, at least 450 mg, or at least 500 mg, of antibody or fragment binding to the antigen thereof. The lyophilized formulation disclosed herein comprising an antibody should be stored at 2 to 8 ° C in its original container and the antibody should be administered within 12 hours, preferably within 6 hours, within 5 hours. hours, within 3 hours, or within 1 hour after being reconstituted. The formulation disclosed herein comprising antibodies can be formulated as neutral or salt forms. Pharmaceutically acceptable salts include those formed with anions such as those derived from hydrochloric, phosphoric, acetic, oxalic, tartaric acids, etc., and those formed with cations such as those derived from sodium, potassium, ammonium, calcium, ferric, isopropylamine hydroxides. , triethylamine, 2-ethylaminoethanol, histidine, procaine, etc.

In a twenty-fourth aspect, the present disclosure relates to a manufacturing article comprising the pharmaceutical composition disclosed herein, the liquid formulation disclosed herein, or the dry formulation disclosed herein, the antibody or fragment of antigen binding thereof disclosed herein, or one or more unit dosage forms disclosed herein and a container or container.

According to another preferred embodiment, the article of manufacture comprises sufficient unit dosage forms of antibody for a period of two weeks (14 days), four weeks (28 days) or one month, with either E2W, E4W or an administration schedule of once a month.

The article of manufacture may comprise one or more unit dosage forms containing both the antibody and the HMG CoA inhibitor, for example, a unit dosage form comprising a liquid formulation for injection or infusion comprising both active ingredients. The article of manufacture may also comprise the antibody (or antigen binding fragment thereof) and the HMG-CoA reductase inhibitor in two or more separate unit dosage forms.

According to one embodiment, the article of manufacture comprises one or more unit dosage forms separated from and the HMG-CoA reductase inhibitor disclosed herein.

According to a preferred embodiment, each unit dosage form of the HMG-CoA reductase inhibitor comprises about 0.05 mg to about 100 mg of HMG-CoA reductase inhibitor.

According to another preferred embodiment, the HMG-CoA reductase inhibitor is a statin, preferably selected from the list containing: cerivastatin, atorvastatin, simvastatin, pitavastatin, rosuvastatin, fluvastatin, lovastatin or pravastatin and preferably atorvastatin.

According to another preferred embodiment, the HMG-CoA reductase inhibitor, for example, statin, is in an effective dose for administration once a day.

According to another preferred embodiment, the unit dosage form of HMG-CoA reductase inhibitor comprises about 0.5 to about 100 mg, about 5 to about 90 mg, of about 10, 20, 40 or 80 mg of HMG-CoA inhibitor reductase

According to another preferred embodiment, the article of manufacture comprises sufficient unit dosage forms of HMG-CoA reductase inhibitor for a daily administration schedule.

According to another preferred embodiment, the unit dosage form comprising the antibody is an envelope, a pre-filled syringe, a pre-filled auto-injector or a cartridge for a reusable syringe or applicator, especially comprising 1 ml or 2 ml of solution for injection.

According to another embodiment, the article of manufacture comprises one or more of the following components:

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i. One or more unit dosage forms comprising the antibody disclosed herein.

j. One or more unit dosage forms comprising the HMG-CoA reductase inhibitor disclosed herein;

k. Instructions for use;

l. A device for administration of the antibody such as a syringe.

According to another preferred embodiment, the article of manufacture comprises sufficient unit dosage forms of the antibody and preferably also of the HMG-CoA reductase inhibitor ...

(a) for a single administration of HMG-CoA reductase antibody and inhibitor, for example, comprising a vial, envelope, vial, cartridge or pre-filled syringe comprising approximately 50 mg, approximately 100 mg, approximately 150 mg, approximately 200 mg , about 250 mg, about 300 mg, about 350 mg, about 400 mg, about 450 mg or about 500 mg of antibody and preferably about 150 mg of antibody, about 200 mg of antibody or about 300 mg of antibody, together with tablet or capsule, for example, for oral or peroral administration comprising the HMG-CoA inhibitor, for example, comprising about 10 mg, about 20 mg, about 40 mg or about 80 mg of the HMG-CoA inhibitor such as atorvastatin.

(b) for a two-week treatment (i.e. 14 days) with the HMG-CoA reductase antibody and inhibitor, for example, comprising a vial, envelope, vial, cartridge or pre-filled syringe comprising approximately 50 mg, approximately 100 mg, approximately 150 mg, approximately 200 mg, approximately 250 mg, approximately 300 mg, approximately 350 mg, approximately 400 mg, approximately 450 mg or approximately 500 mg of antibody and preferably approximately 150 mg of antibody, approximately 200 mg of antibody or approximately 300 mg of antibody; together with sufficient units comprising of HMG-CoA reductase inhibitor (for example, tablets or capsules, for example, for oral or peroral administration) for a 14-day treatment, for example, 14 units for a one-time administration schedule per day of HMG-CoA reductase inhibitor or 28 units for a twice daily administration schedule, etc., wherein the units per day of HMG-CoA inhibitor preferably comprise approximately 10 mg, approximately 20 mg, approximately 40 mg or about 80 mg of the HMG-CoA inhibitor such as atorvastatin. In the event that the antibody is to be administered in a dosage greater than 200 mg, two unit dosage forms of antibody together comprising the total dose may be preferable (for example, two pre-filled syringes comprising approximately 150 mg of antibody in 1 ml of liquid formulation each for a total administration (eg, subcutaneous injection) of approximately 300 mg of antibody in two injections) (or two units with approximately 100 mg each for a total administration of approximately 200 mg of antibody, two units with approximately 175 mg for a total administration of approximately 350 antibody, etc.).

(c) for a four-week treatment (i.e. 28 days) with antibody and HMG-CoA reductase inhibitor, for example,

1. for an E2W administration schedule of the antibody with about 50 to about 200 mg of antibody for two weeks: comprising two unit dosage forms (eg, as exemplified above) with each of about 50 mg, about 100 mg , approximately 150 mg or approximately 200 mg of antibody or antigen binding fragment thereof together with 28 unit dosage forms of HMG-CoA reductase inhibitor (as exemplified above) for a once-daily daily administration schedule or together with 56 unit dosage forms of HMG-CoA reductase inhibitor for a twice daily daily administration schedule, preferably 28 unit dosage forms (eg, capsules or tablets) of approximately 10 mg, approximately 20 mg, approximately 40 mg or about 80 mg atorvastatin

2. for an E4W administration schedule of the antibody or fragment thereof with an administration of approximately 200 mg for four weeks (28 days): for example, comprising a unit dosage form of the antibody with approximately 200 mg of antibody (for example , as exemplified above) together with 28 or 56, and preferably 28 unit dosage forms of HMG-CoA reductase inhibitor (for example, as exemplified above)

3. for an E4W administration regimen of the antibody with more than 200 mg for four weeks (28 days): comprising two unit dosage forms that together comprise the total antibody dose (eg, two pre-filled syringes each comprising 1 ml of liquid antibody formulation with 150 mg of antibody each) or comprising a unit dosage form comprising the total amount of antibody to be administered (eg, a vial that

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comprises approximately 300 mg of antibody for dissolution or a pre-filled vial, cartridge or syringe comprising approximately 300 mg of the antibody in liquid formulation (i.e., approximately 2 ml of liquid formulation, wherein 1 ml of liquid formulation comprises approximately 150 mg of the antibody); together with 28 or 56, and preferably 28 unit dosage forms of HMG-CoA reductase inhibitor (for example, as exemplified above)

(d) for a one month treatment with antibody and HMG-CoA reductase inhibitor: comprising the same numbers of unit dosage forms of antibody as exemplified in (c) for once or twice monthly administration, for example, every first day of the month or every first Monday, etc., of the month for a once-a-month administration, or for example, every first and 14th or 15th day of the month for a twice-daily administration schedule month; furthermore, the article of manufacture comprises 31 unit dosage forms of HMG-CoA reductase inhibitor, preferably tablets or capsules arranged in a honeycomb container containing a consecutive numbering of 1-31 for the days of the month (in which the tablets or Superfluous capsules for excess days should be discarded).

In a twenty-fifth aspect, the present disclosure refers to an article of manufacture comprising: (a) a packaging material; (b) an antibody or an antigen binding fragment thereof that specifically binds to PCSK9h; and (c) a label or leaflet contained within the packaging material that indicates that patients receiving treatment with said antibody or antigen binding fragment can be treated for a disease or condition selected from the group consisting of hypercholesterolemia, hyperlipidemia, dyslipidemia, Atherosclerosis and cardiovascular diseases and additionally indicating that subjects that are in one or more groups of subjects can be treated as cited in the thirteenth aspect.

In a twenty-sixth aspect, the present disclosure refers to an article of manufacture comprising: (a) a packaging material; (b) an antibody or an antigen binding fragment thereof that specifically binds to PCSK9h; and (c) a label or leaflet contained within the packaging material that indicates that patients receiving treatment with said antibody or antigen binding fragment can be treated for a disease or condition selected from the group consisting of hypercholesterolemia, hyperlipidemia, dyslipidemia, Atherosclerosis and cardiovascular diseases and additionally indicating that the treatment of patients with said antibody or antigen binding fragment thereof is contraindicated for patients belonging to one or more groups of subjects as cited in the fourteenth aspect.

Antibodies and antigen-binding fragments thereof that can be used to practice the different articles of manufacture disclosed herein are described in the "Preferred antibodies" section.

In preferred embodiments of the twenty-fourth, the twenty-fifth or twenty-sixth aspect, the label or package leaflet contains a reference to a method of treatment according to the first or medical uses according to the second aspect and the embodiments of the first and second aspects as described in This document.

The articles of manufacture described in the 24th, the 25th or the 26th aspect may further comprise one or more of the features or components of the manufacturing article as comprised in the 3rd, 4th or 5th aspect disclosed in the present document and vice versa. A further preferred embodiment of the present disclosure combines one or more of the characteristics of 24 ° and 25 °, 25 ° and 26 ° or 24 ° and 25 ° and 26 ° aspect as described herein. .

According to a 27th aspect, the present disclosure refers to a pharmaceutical composition or antibody or antigen binding fragment thereof disclosed herein, as per the ninth aspect of the present disclosure, for use in the treatment of a disease or condition in which the expression or activity of PCSK9 causes an impact, preferably for use in reducing elevated levels of LDL-C (low density lipoprotein C).

According to a preferred embodiment, the disease or condition is selected from the group consisting of: elevated levels of total cholesterol, elevated levels of low density lipoprotein (LDL-C), hypercholesterolemia, hyperlipidemia, dyslipidemia and atherosclerosis, particularly primary hypercholesterolemia, familial hypercholesterolemia , or hypercholesterolemia that is not controlled by statins

According to another preferred embodiment, the composition, the antibody or antigen binding fragment thereof is administered to the subject every two weeks (E2W), every four weeks (E4W) or once a month.

According to another preferred embodiment, an HMG-CoA reductase inhibitor is co-administered with the pharmaceutical composition, the antibody or antigen binding fragment thereof, preferably an HMG-CoA reductase inhibitor according to one of the different aspects disclosed herein. document, as per the first or second aspect.

According to another preferred embodiment, the HMG-CoA reductase inhibitor is administered once daily and preferably every day.

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In a twenty-eighth aspect, the present disclosure relates to a method of preparing a pharmaceutical composition disclosed herein, for example, according to the nineteenth aspect, comprising mixing the antibody or antigen-binding fragment thereof and optionally the HMG-CoA reductase inhibitor with one or more pharmaceutical excipients or vehicles.

In a twenty-ninth aspect, the present disclosure relates to a method of preparing a unit dosage form herein comprising mixing an amount of the pharmaceutical composition, of the antibody or antigen-binding fragment thereof, of the liquid formulation or of the dry formulation disclosed herein comprising one or more doses of the antibody or antigen fragment thereof and optionally of the HMG-CoA reductase inhibitor and adapting them as physically discrete units suitable as unit dosages for human and / or animal administration. .

In a thirtieth aspect, the present disclosure relates to a method of preparing or assembling an article of manufacture disclosed herein comprising packaging the pharmaceutical composition, of the antibody according to, of the liquid formulation, of the dry formulation according to or a or more of the unit dosage forms disclosed herein in a container, optionally together with one or more of the following: a label, instructions for use, an administration device.

In a thirty-first aspect, the present disclosure relates to a method of testing the efficacy of an antibody or an antigen binding fragment thereof that specifically binds to PCSK9h for the treatment of a disease or condition selected from the group consisting of in elevated levels of LDL-C, hypercholesterolemia, hyperlipidemia, dyslipidemia, atherosclerosis and cardiovascular diseases, or any other disease or condition described in the first or second aspect, said method comprising:

treating a selected patient population with said antibody or antigen binding fragment thereof, in which each patient in said population has a cholesterol level of (C-LDL) greater than 100 mg / dl; Y

determining the efficacy of said antibody or antigen binding fragment thereof by determining the level of LDL-C in the patient population before and after administration of said antibody or antigen binding fragment thereof, in which a reduction in the LDL-C level of at least 25% with respect to a predose level in at least 75% of the patient population indicates that said antibody or antigen-binding fragment thereof is effective for the treatment of said disease or condition in said patient population;

in which each patient is classified into one or more groups of subjects as cited in the thirteenth aspect.

In a thirty-second aspect, the present disclosure relates to a method of testing the efficacy of an antibody or an antigen binding fragment thereof that specifically binds to PCSK9h for the treatment of a disease or condition selected from the group consisting of in elevated levels of LDL-C, hypercholesterolemia, hyperlipidemia, dyslipidemia, atherosclerosis and cardiovascular diseases (or any other method as described in the first or second aspect), said method comprising:

determining the efficacy of an antibody or antigen binding fragment thereof that has been used for the treatment of a selected patient population with said antibody or antigen binding fragment thereof, in which each patient in said population has a level of LDL cholesterol greater than 100 mg / dl determining the level of LDL-C in the patient population before and after the administration of said antibody or antigen-binding fragment thereof, in which a reduction in the level of C- LDL of at least 25% with respect to a predose level in at least 75% of the patient population indicates that said antibody or antigen-binding fragment thereof is effective for the treatment of said disease or condition in said population of patients;

in which each patient is classified into one or more groups of subjects as cited in the thirteenth aspect.

In preferred embodiments of the 31st or 32nd aspect, each patient in said population has received a lipid-lowering treatment by administration of an HMG-CoA reductase inhibitor such as a statin for at least 6 weeks before treatment with said antibody or binding fragment. to the antigen thereof.

Antibodies and antigen-binding fragments thereof that can be used to practice the tenth and twentieth aspects of the present disclosure are described in the "Preferred antibodies" section or another section of the present application describing antibodies of the present disclosure. , such as, for example, the fifteenth aspect.

In preferred embodiments of the 31st or 32nd aspect, the selected patient population is or has been treated with a treatment method according to the first or second aspect and the embodiments of the first or second aspect as described herein.

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In further preferred embodiments of the 31st or 32nd aspect, the efficacy of said antibody or said antigen binding fragment thereof is determined for subgroups of said selected patient population, in which said subgroups have been stratified from at least a stratification factor selected from the group consisting of: population with heterozygous familial hypercholesterolemia (heFH); previous history of myocardial infarction (MI); previous history of stroke; who receives high intensity statin therapy; and geographic region of the patient (for example, North America, Western Europe, Eastern Europe and the rest of the world).

In hamsters and other rodents, statins are not effective in removing LDL from the blood. More specifically, the administration of statins alone (for example, atorvastatin) has no effect on the expression of the LDL receptor (LDLR) in hamsters or other rodents, supposedly due to the activity of endogenous PCSK9. The experiments contained in the present application (see study 4) show that inhibition of PCSK9 by administration of an anti-PCSK9 antibody makes rodents (eg, hamsters) sensitive to statin treatment. Therefore, the present application provides a new animal model for testing the efficacy of statins or other drugs that reduce cholesterol levels.

Thus, in a thirty-third aspect, the present disclosure refers to a method of testing the efficacy of a compound in reducing cholesterol levels in a subject, comprising the steps:

(a) provide a rodent;

(b) administering an antibody or an antigen binding fragment thereof that specifically binds PCSK9 to the rodent;

(c) administering a test compound to said rodent;

(d) determine the effect of the test compound on the rodent, in which a reduction in the level of cholesterol in the rodent compared to the cholesterol level of a control animal indicates that the test compound is effective in reducing the levels of cholesterol in a subject, in which the control animal is of the same species as said rodent, and in which the control animal has not been exposed to the test compound.

In preferred embodiments of the thirty-third aspect, the rodent is selected from the group consisting of hamster, mouse, rat, guinea pig and rabbit.

Antibodies and antigen-binding fragments thereof that can be used to implement the twenty-first aspect of the present disclosure are described in the other sections of the present application such as the fifteenth aspect in the "Preferred antibodies" section. Preferably, the antibody or antigen binding fragment thereof is administered to the rodent in a concentration of 1 mg / kg body weight, 3 mg / kg body weight, or 10 mg / kg body weight.

In preferred embodiments of the 33rd aspect, the reduction in cholesterol level is determined by measuring the level of total cholesterol in the serum. In more preferred embodiments, the reduction of the cholesterol level is determined by measuring the level of LDL (C-LDL) cholesterol in the serum.

In preferred embodiments of the 33rd aspect, the control animal is of the same strain as the rodent. Preferably, the same antibody or antigen binding fragment thereof is administered to the rodent and the control animal. Preferably, the same concentration (measured in mg / kg body weight) of the antibody or antigen binding fragment thereof is administered to the rodent and the control animal.

In an embodiment of the 33rd aspect, the control animal is a different animal, that is, a different individual, from the rodent. It is also possible to determine the level of cholesterol in two or more control animals and calculate the average value of the level of cholesterol in these two or more control animals. Likewise, it is possible to expose two or more rodents to the antibody or antigen binding fragment thereof, to determine the level of cholesterol in these two or more rodents and to calculate the average value of the level of cholesterol in these two or more rodents.

In an alternative embodiment of the 33rd aspect, the control animal is the same animal as the rodent but is examined at a different time. More specifically, the level of cholesterol in the rodent after administration of the test compound can be compared with a pre-dose cholesterol level in the same animal. Preferably, said pre-dose cholesterol level is determined between steps (b) and (c) cited above.

According to the thirty-fourth aspect, the present disclosure refers to a method of enhancing the activity of LDL-C reduction in a subject receiving statin therapy, the method comprising administering to the subject an antibody, or antigen-binding fragment thereof, which specifically binds to proprotein convertase subtilisin / human type 9 kexin (PCSK9h), in which the antibody or antigen binding fragment thereof is administered in a dose amount within the range of about 5 mg to about 500 mg, thereby enhancing the C-LCL reducing activity of statin therapy in the subject.

According to a preferred embodiment of the 34th aspect, the subject is resistant to statin therapy before antibody administration.

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According to another preferred embodiment, the subject suffers from a disease or condition selected from the group consisting of hypercholesterolemia, hyperlipidemia, dyslipidemia and atherosclerosis.

According to another preferred embodiment, the pathology is primary hypercholesterolemia or familial hypercholesterolemia.

According to another preferred embodiment, the antibody or antigen binding fragment is administered in a dose amount within the range of about 50 mg to about 300 mg.

According to another preferred embodiment, the antibody or antigen binding fragment is administered in a dose amount of approximately 150 mg.

According to another preferred embodiment, the antibody or antigen binding fragment thereof is administered to the subject every two weeks (E2W).

According to another preferred embodiment, the antibody or antigen binding fragment thereof is administered to the subject every four weeks (E4W).

According to another preferred embodiment, the treatment reduces total serum cholesterol at least about 25% to about 35% with respect to a predose level and maintains the reduction for at least a 24-day period.

According to another preferred embodiment, the treatment reduces total serum cholesterol at least about 65% to about 80% with respect to a predose level and maintains the reduction for at least a 24-day period.

According to another preferred embodiment, the treatment reduces serum triglyceride levels from at least about 25% to about 40% with respect to a predose level.

According to another preferred embodiment, the treatment reduced serum HDL cholesterol no more than 5% with respect to a predose level.

According to another preferred embodiment, the treatment has little or no measurable effect on liver function, as determined by measurements of ALT and AST.

According to another preferred embodiment, the antibody or antigen binding fragment comprises the heavy and light chain CDRs of a pair of HCVR / LCVR amino acid sequences as shown in SEQ ID NO: 90/92.

According to another preferred embodiment, the antibody or antigen binding fragment comprises a pair of amino acid sequences of HCVR / LCVR as shown in SEQ ID NO: 90/92.

According to another preferred embodiment, the antibody or antigen binding fragment thereof competes to bind PCSK9h with an antibody or antigen binding fragment comprising a pair of amino acid sequences of HCVR / LCVR as shown in SEQ ID NO: 90 / 92.

According to another preferred embodiment, the statin is selected from the group consisting of cerivastatin, atorvastatin, simvastatin, pitavastatin, rosuvastatin, fluvastatin, lovastatin and pravastatin.

According to another preferred embodiment, the statin is atorvastatin administered at a dose of 10 mg, 20 mg, 40 mg or 80 mg.

In a thirty-fifth aspect, the present disclosure relates to a kit for treating high levels of low density lipoprotein (C-LDL) cholesterol in a subject, the kit comprising (a) unit dosage pharmaceutical form comprising an antibody, or antigen binding fragment thereof, which specifically binds to PCSK9h; and pharmaceutically acceptable carrier, wherein the antibody or antigen-binding fragment is present in a dose amount within the range of about 5 mg to about 500 mg; and (b) a label or leaflet with instructions for its use.

According to a preferred embodiment of the 35th aspect, the label indicates that patients receiving treatment with said antibody or antigen binding fragment can be treated for a disease or condition selected from the group consisting of hypercholesterolemia, hyperlipidemia, dyslipidemia and atherosclerosis and cardiovascular diseases. .

According to another preferred embodiment, the disease or condition is primary hypercholesterolemia or familial hypercholesterolemia. According to another preferred embodiment, the disease or condition is hypercholesterolemia that is not controlled by statins.

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According to another preferred embodiment, the antibody or antigen binding fragment is present in a dose amount within the range of about 50 mg to about 300 mg. According to another preferred embodiment, the antibody or antigen binding fragment is present in a dose amount of approximately 150 mg.

According to another preferred embodiment, the label or leaflet indicates that the antibody or antigen binding fragment thereof is administered to the subject every two weeks (E2W).

According to another preferred embodiment, the label or leaflet indicates that the antibody or antigen binding fragment thereof is administered to the subject every four weeks (E4W).

According to another preferred embodiment, the antibody or antigen binding fragment comprises the heavy and light chain CDRs of a pair of HCVR / LCVR amino acid sequences as shown in SEQ ID NO: 90/92

According to another preferred embodiment, the antibody or antigen binding fragment comprises a pair of amino acid sequences of HCVR / LCVR as shown in SEQ ID NO: 90/92.

According to another preferred embodiment, the antibody or antigen binding fragment thereof competes to bind PCSK9h with an antibody or antigen binding fragment comprising a pair of amino acid sequences of HCVR / LCVR as shown in SEQ ID NO: 90 / 92.

According to another preferred embodiment, the kit further comprises an HMG-CoA reductase inhibitor. According to another preferred embodiment, the inhibitor is in a dose amount in the range of about 0.05 mg to 100 mg. According to another preferred embodiment, the HMG-CoA reductase inhibitor is a statin. According to another preferred embodiment, the statin is selected from the group consisting of cerivastatin, atorvastatin, simvastatin, pitavastatin, rosuvastatin, fluvastatin, lovastatin and pravastatin.

According to another preferred embodiment, the instructions indicate that the statin is atorvastatin administered at a dose of 10 mg, 20 mg, 40 mg or 80 mg.

According to another preferred embodiment, the instructions indicate that treatment with the antibody or one is contraindicated for patients belonging to one or more of the following groups:

(i) smokers;

(ii) persons 70 years of age or older;

(iii) people suffering from hypertension;

(iv) women who are pregnant;

(v) women who are trying to get pregnant;

(vi) women who are breastfeeding;

(vii) people who have or have ever had a disease that affects the liver;

(viii) people who had some unexplained abnormal blood test for liver function;

(ix) people who drink excessive amounts of alcohol;

(x) people who have kidney problems;

(xi) people suffering from hypothyroidism;

(xii) people suffering from muscular disorders;

(xiii) people who have had previous muscle problems during treatment with lipid lowering medicine;

(xiv) people who have serious problems with their breathing;

(xv) people taking one or more of the following medicines: medicines that alter the way the immune systems work (for example, cyclosporine or antihistamines), antibiotics or antifungal medicines (for example, erythromycin, clarithromycin, ketoconazole, itraconazole , rifampicin, fusidic acid), medicines that regulate lipid levels (for example, gemfibrozil, colestipol), calcium channel blockers (for example, verapamil, diltiazem), medicines that regulate heart rhythm (digoxin, amiodarone), protease inhibitors used in the treatment of HIV (for example, nelfinavir), warfarin, oral contraceptives, antacids or St. John's wort; or

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(xvi) people who drink more than 0.1 l of grapefruit juice a day;

(xvii) people who have a body mass index (BMI) greater than 40;

(xviii) people who have a body mass index (BMI) of less than 18;

(xix) people suffering from type 1 diabetes or type 2 diabetes;

(xx) people positive for hepatitis B or hepatitis C; or

(xxi) people who have a known sensitivity to therapeutic drugs of monoclonal antibodies.

In a thirty-sixth aspect, the present disclosure refers to a method of treating a subject suffering from a disease or disorder characterized by elevated levels of low density lipoprotein cholesterol (LDL-C), the method comprising:

(a) select a subject with a blood LDL-C level greater than 100 mg / dl; Y

(b) administering to said subject a composition comprising an antibody or antigen binding fragment thereof that specifically binds to proprotein convertase subtilisin / human type 9 kexin (PCSK9h); thus reducing cholesterol levels in the subject in need thereof.

According to a preferred embodiment, the disease or condition is selected from the group consisting of: hypercholesterolemia, hyperlipidemia, dyslipidemia and atherosclerosis.

According to another preferred embodiment, the pathology is primary hypercholesterolemia or familial hypercholesterolemia.

According to another preferred embodiment, the disease or condition is hypercholesterolemia that is not controlled by statins.

According to another preferred embodiment, the subject has a body mass index (BMI) of less than 18 kg / m2 or greater than 40 kg / m2

According to another preferred embodiment, the subject was not previously informed to participate in a hypocholesterolemic diet.

According to another preferred embodiment, the subject has not previously taken a hypocholesterolemiant drug except atorvastatin.

According to another preferred embodiment, atorvastatin was administered at approximately 10 mg per day.

According to another preferred embodiment, the hypocholesterolemiant drug is selected from the group consisting of fibrates, bile acid resins, niacin, intestinal cholesterol absorption blockers (ICA) and omega-3 fatty acids. According to another preferred embodiment, niacin is administered at more than 500 mg per day. According to another preferred embodiment, omega-3 fatty acids are administered at more than 1000 mg per day.

According to another preferred embodiment, the subject does not suffer from diabetes. According to another preferred embodiment, the diabetes is type 1 diabetes. According to another preferred embodiment, the diabetes is type 2 diabetes. According to another preferred embodiment, type 2 diabetes is treated with insulin.

According to another preferred embodiment, the subject has a glycosylated hemoglobin concentration in blood greater than or equal to 8.5%.

According to another preferred embodiment, the subject is negative for hepatitis B and surface antigen C.

According to another preferred embodiment, the subject has a blood triglyceride concentration greater than 350 mg / dl.

According to another preferred embodiment, the subject has less than 1500 neutrophils per cubic mm of blood.

According to another preferred embodiment, the subject has less than 100,000 platelets per cubic mm of blood.

According to another preferred embodiment, the subject is a female.

According to another preferred embodiment, the subject is not pregnant.

According to another preferred embodiment, the subject has a concentration of thyroid stimulating hormone in the blood that is above the lower limit of normal and below the upper limit of normal.

According to another preferred embodiment, the subject has serum creatine less than 1.4 of the upper limit of normal.

According to another preferred embodiment, the subject is a male.

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According to another preferred embodiment, the subject has serum creatine less than 1.5 of the upper limit of normal.

According to another preferred embodiment, the subject has an amount of aspartate transaminase that is less than twice the upper limit of normal.

According to another preferred embodiment, the subject has an amount of alanine transaminase that is less than twice the upper limit of normal.

According to another preferred embodiment, the antibody or antigen binding fragment is administered in a dose amount within the range of about 5 mg to about 500 mg.

According to another preferred embodiment, the antibody or antigen binding fragment is administered in a dose amount within the range of about 50 mg to about 300 mg.

According to another preferred embodiment, the antibody is administered at 200 to 300 mg every four weeks.

According to another preferred embodiment, the antibody or antigen binding fragment is administered in a dose amount of approximately 150 mg.

According to another preferred embodiment, the antibody or antigen binding fragment thereof is administered to the subject every two weeks (E2W).

According to another preferred embodiment, the antibody or antigen binding fragment thereof is administered to the subject every four weeks (E4W).

According to another preferred embodiment, the antibody or antigen binding fragment comprises the heavy and light chain CDRs of a pair of HCVR / LCVR amino acid sequences as shown in SEQ ID NO: 90/92.

According to another preferred embodiment, the antibody or antigen binding fragment comprises a pair of amino acid sequences of HCVR / LCVR as shown in SEQ ID NO: 90/92.

According to another preferred embodiment, the antibody or antigen binding fragment thereof competes to bind PCSK9h with an antibody or antigen binding fragment comprising a pair of amino acid sequences of HCVR / LCVR as shown in SEQ ID NO: 90 / 92. According to another preferred embodiment, the antibody is administered subcutaneously. According to another preferred embodiment, the antibody is administered in the abdomen.

According to another preferred embodiment, an HMG-CoA reductase inhibitor is administered to the subject.

According to another preferred embodiment, the HMG-CoA reductase inhibitor is administered in a dose amount in the range of about 0.05 mg to 100 mg.

According to another preferred embodiment, the HMG-CoA reductase inhibitor is a statin.

According to another preferred embodiment, the statin is selected from the group consisting of cerivastatin, atorvastatin, simvastatin, pitavastatin, rosuvastatin, fluvastatin, lovastatin and pravastatin.

According to another preferred embodiment, the statin is atorvastatin administered at a dose of 10 mg or 80 mg.

According to another preferred embodiment, atorvastatin is administered at about 10 mg per day and at 80 mg one day in a period of 8 weeks.

In a thirty-seventh aspect, the present disclosure refers to a method of reducing cholesterol levels in a subject in need thereof, comprising:

(a) select a subject with a low density lipoprotein cholesterol (C-LDL) level in blood greater than 100 mg / dl; Y

(b) administering to said subject a composition comprising an antibody or antigen binding fragment thereof that specifically binds to proprotein convertase subtilisin / human type 9 kexin (PCSK9h); thus reducing cholesterol levels in the subject in need thereof.

According to a preferred embodiment of the 37th aspect, the disease or condition is selected from the group consisting of: hypercholesterolemia, hyperlipidemia, dyslipidemia and atherosclerosis.

According to another preferred embodiment, the pathology is primary hypercholesterolemia or familial hypercholesterolemia.

According to another preferred embodiment, the disease or condition is hypercholesterolemia that is not controlled by statins.

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According to another preferred embodiment, the subject has a body mass index (BMI) of less than 18 kg / m2 or greater than 40 kg / m2

According to another preferred embodiment, the subject was not previously informed to participate in a hypocholesterolemic diet.

According to another preferred embodiment, the subject has not previously taken a hypocholesterolemiant drug except atorvastatin.

According to another preferred embodiment, atorvastatin was administered at approximately 10 mg per day.

According to another preferred embodiment, the hypocholesterolemiant drug is selected from the group consisting of fibrates, bile acid resins, niacin, intestinal cholesterol absorption blockers (ICA) and omega-3 fatty acids.

According to another preferred embodiment, niacin is administered at more than 500 mg per day.

According to another preferred embodiment, omega-3 fatty acids are administered at more than 1000 mg per day.

According to another preferred embodiment, the subject does not suffer from diabetes.

According to another preferred embodiment, the diabetes is type 1 diabetes.

According to another preferred embodiment, the diabetes is type 2 diabetes.

According to another preferred embodiment, type 2 diabetes is treated with insulin.

According to another preferred embodiment, the subject has a higher blood glycosylated hemoglobin concentration or

equal to 8.5%.

According to another preferred embodiment, the subject is negative for hepatitis B and surface antigen C.

According to another preferred embodiment, the subject has a blood triglyceride concentration greater than 350 mg / dl. According to another preferred embodiment, the subject has less than 1500 neutrophils per cubic mm of blood.

According to another preferred embodiment, the subject has less than 100,000 platelets per cubic mm of blood.

According to another preferred embodiment, the subject is a female.

According to another preferred embodiment, the subject is not pregnant.

According to another preferred embodiment, the subject has a concentration of thyroid stimulating hormone in the blood that is above the lower limit of normal and below the upper limit of normal.

According to another preferred embodiment, the subject has serum creatine less than 1.4 of the upper limit of normal.

According to another preferred embodiment, the subject is a male.

According to another preferred embodiment, the subject has serum creatine less than 1.5 of the upper limit of normal.

According to another preferred embodiment, the subject has an amount of aspartate transaminase that is less than twice the upper limit of normal.

According to another preferred embodiment, the subject has an amount of alanine transaminase that is less than twice the upper limit of normal.

According to another preferred embodiment, the antibody or antigen binding fragment is administered in a dose amount within the range of about 5 mg to about 500 mg.

According to another preferred embodiment, the antibody or antigen binding fragment is administered in a dose amount within the range of about 50 mg to about 300 mg.

According to another preferred embodiment, the antibody is administered at 200 to 300 mg every four weeks.

According to another preferred embodiment, the antibody or antigen binding fragment is administered in a dose amount of approximately 150 mg.

According to another preferred embodiment, the antibody or antigen binding fragment thereof is administered to the subject every two weeks (E2W).

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According to another preferred embodiment, the antibody or antigen binding fragment thereof is administered to the subject every four weeks (E4W).

According to another preferred embodiment, the antibody or antigen binding fragment comprises the heavy and light chain CDRs of a pair of HCVR / LCVR amino acid sequences as shown in SEQ ID NO: 90/92. According to another preferred embodiment, the antibody or antigen binding fragment comprises a pair of amino acid sequences of HCVR / LCVR as shown in SEQ ID NO: 90/92. According to another preferred embodiment, the antibody or antigen binding fragment thereof competes to bind PCSK9h with an antibody or antigen binding fragment comprising a pair of amino acid sequences of HCVR / LCVR as shown in SEQ ID NO: 90 / 92. According to another preferred embodiment, the antibody is administered subcutaneously. According to another preferred embodiment, the antibody is administered in the abdomen.

According to another preferred embodiment, the method further comprises administering an HMG-CoA reductase inhibitor to the subject. According to another preferred embodiment, the HMG-CoA reductase inhibitor is administered in a dose amount in the range of about 0.05 mg to 100 mg. According to another preferred embodiment, the HMG-CoA reductase inhibitor is a statin. According to another preferred embodiment, the statin is selected from the group consisting of cerivastatin, atorvastatin, simvastatin, pitavastatin, rosuvastatin, fluvastatin, lovastatin and pravastatin. According to another preferred embodiment, the statin is atorvastatin administered at a dose of 10 mg, 20 mg, 40 mg or 80 mg. According to another preferred embodiment, atorvastatin is administered at about 10 mg per day and at 80 mg one day in a period of 8 weeks.

Several aspects of the disclosure can be combined with each other. For example, the method of treatment of a disease or condition can be combined according to the first aspect and the method of treatment of a disease or condition according to the thirteenth aspect. As a result of this combination, the present disclosure refers to a method of treatment of a disease or condition that characterizes the treatment of certain groups of subjects by certain dosage guidelines. Similarly, the antibody or antigen-binding fragment for use in the treatment of a disease or condition according to the second aspect may be combined with the antibody or antigen-binding fragment for use in the treatment of a disease or condition. according to the fifteenth aspect. As a result of this combination, the present disclosure relates to an antibody or antigen binding fragment thereof for use in the treatment of certain groups of subjects by certain dosage guidelines.

According to another example, the method of treatment of a disease or condition can be combined according to the first aspect and the method of treatment of a disease or condition according to the fourteenth aspect. As a result of this combination, the present disclosure refers to a method of treating a disease or condition that excludes certain groups of subjects from a treatment by a certain dosage schedule. Similarly, the antibody or antigen-binding fragment for use in the treatment of a disease or condition according to the second aspect may be combined with the antibody or antigen-binding fragment for use in the treatment of a disease or condition. according to the sixteenth aspect. As a result of this combination, the present disclosure refers to an antibody or antigen binding fragment thereof for use in the treatment by a certain dosage schedule, in which certain groups of subjects are excluded from the treatment.

The expert will recognize other preferred embodiments resulting from suitable combinations of different aspects and embodiments of the present disclosure.

The pharmaceutical uses of the present disclosure as described herein also refer to uses of a given antibody or antigen binding fragment thereof, of the given pharmaceutical composition, etc., for the manufacture of a medicament for treatment. of one or more of the diseases or conditions as described herein.

Preferred Antibodies

The following section describes functional and structural characteristics of antibodies and antigen binding fragments thereof that can be used to practice the twenty-one aspects of the present disclosure. Thus, expressions such as "in preferred embodiments", "in some embodiments", "in another preferred embodiment" and similar expressions should be understood as referring to embodiments of the first aspect of the present disclosure, the second aspect of the present disclosure, the third aspect of the present disclosure, the fourth aspect of the present disclosure, the fifth aspect of the present disclosure, the sixth aspect of the present disclosure, the seventh aspect of the present disclosure, the eighth aspect of the present disclosure, the ninth aspect of the present disclosure, the tenth aspect of the present disclosure, the eleventh aspect of the present disclosure, the twelfth aspect of the present disclosure, the thirteenth aspect of the present disclosure, the fourteenth aspect of the present disclosure, the 15th aspect of this

disclosure, the sixteenth aspect of this disclosure, the seventeenth aspect of this

disclosure, the eighteenth aspect of the present disclosure, the eighteenth aspect of the present

disclosure, the twenty-first aspect and the twenty-first aspect of the present disclosure, the twenty-second

aspect of the present disclosure, the twenty-third aspect of the present disclosure, the twenty-fourth aspect of the present disclosure, the twenty-fifth aspect of the present disclosure, the twenty-sixth aspect of the

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present disclosure, the twenty-seventh aspect of the present disclosure, the twenty-eighth aspect of the present disclosure, the twenty-ninth aspect of the present disclosure, the thirtieth aspect of the present disclosure

disclosure, the thirty-first aspect of the present disclosure, the thirty-second aspect of the present

disclosure, the thirty-third aspect of the present disclosure, the thirty-fourth aspect of the present

disclosure, the thirty-fifth aspect of the present disclosure, the thirty-sixth aspect of the present

disclosure, the thirty-seventh aspect of the present disclosure.

All antibodies or antigen-binding fragments thereof suitable for practicing the present disclosure specifically bind PCSK9h. In preferred embodiments of any aspect of the present disclosure, the antibody or antigen binding fragment thereof is a recombinant human antibody or fragment thereof. In more specific embodiments, the antibody or antigen binding fragment thereof is a fully human monoclonal antibody or antigen binding fragment thereof that specifically binds PCSK9h and neutralizes the activity of PCSK9.

The mAbs usable in the present disclosure may be full length (for example, an IgG1 or IgG4 antibody) or may comprise only an antigen binding portion (eg, a Fab, F (ab ') 2 or scFv fragment), and can be modified to affect functionality, for example, to eliminate residual effector functions (Reddy et al. (2000) J. Immunol. 164: 1925-1933).

In preferred embodiments, the antibodies of the present disclosure are characterized by one or more of the following characteristics after administration to a subject, preferably a human or non-human mammal and more preferably a human:

- Reduction of low-density lipoprotein-C (C-LDL) levels of at least about -25% at

approximately -40% with respect to a predose level with a sustained reduction for at least one

14 day period, in which the sustained reduction is preferably at least -25% and more preferably at least -30% with respect to a predose level, particularly if administered in a dose of about 40 to about 60 mg, preferably about 45 to about 55 mg and more preferably about 50 mg in a biweekly administration schedule (every two weeks, E2W),

- Reduction of low-density lipoprotein-C (C-LDL) of at least about -50% at

approximately -65% with respect to a predose level with a sustained reduction for at least one

14 day period, in which the sustained reduction is preferably at least -40% and more preferably at least -45% with respect to a predose level, particularly if administered in a dose of approximately 100 mg E2W,

- reduction of low-density lipoprotein-C (C-LDL) from at least about -60% to at least about -75% [eg, at least about -60%, at least about -65%, at least about -70 or at least about -75%] with respect to a predose level with a sustained reduction for at least a period of 14 days, in which the sustained reduction is preferably at least -55% and more preferably at least -60% with respect to a predose level, particularly when administered in a dose of approximately 150 mg E2W,

- Reduction of low-density lipoprotein-C (C-LDL) of at least about 40% at

approximately 75% with respect to a predose level with a sustained reduction for at least a period of 28 days, in which the sustained reduction is preferably at least -35% and more preferably at least -40% with respect to a predose level, particularly when administered in a dose of approximately 200 mg E4W,

- Reduction of low-density lipoprotein-C (C-LDL) of at least about -50% at

approximately -75% with respect to a predose level with a sustained reduction for at least a period of 28 days, in which the sustained reduction is preferably at least -40% and more preferably at least -45% with respect to a predose level, particularly when administered in a dose of approximately 300 mg E4W,

- increased serum HDL cholesterol levels of at least 2%, at least 2.5%, at least 3%,

at least 3.5%, at least 4%, at least 4.5%, at least 5% or at least 5.5% with respect to a

predose level, particularly when administered at a dose of approximately 150 mg E2W,

- little or no detectable effect on troponin levels,

- increase in one or more of: total cholesterol levels, ApoB levels, C-non HDL levels, Apo-B / ApoA-1 ratio,

The antibodies according to the present disclosure have the above properties preferably if they are administered in combination with a treatment with HMG-CoA reductase inhibitor. Preferred embodiments of the HMG-CoA reductase inhibitors to be used in conjunction with the antibody disclosed herein

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Document and dosage guidelines and administration thereof can be found throughout the specification, particularly as described in aspects related to medical uses and treatment methods.

According to another preferred embodiment of the antibodies and antigen binding fragments thereof disclosed herein, the antibody or antigen binding fragment thereof has one or more of the following characteristics:

- The antibody or antigen binding fragment comprises the heavy and light chain CDRs of a pair of HCVR / LCVR amino acid sequences as shown in SEQ ID NO: 90/92.

- The antibody or antigen binding fragment thereof comprises a pair of HCVR / LCVR amino acid sequences as shown in SEQ ID NO: 90/92.

- The antibody or antigen binding fragment thereof competes to bind PCSK9h with an antibody or antigen binding fragment comprising a pair of amino acid sequences of HCVR / LCVR as shown in SEQ ID NO: 90/92.

According to another preferred embodiment of the antibodies and antigen binding fragments thereof disclosed herein, the antibody or antigen binding fragment thereof has one or more of the following characteristics:

- overcomes statin resistance in mammals, especially in rodents such as hamster

- increase in LDLR expression in mammals, particularly in rodents such as hamster

- decreases serum C-LDL in rodents such as hamster

- synergistic decrease of C-LDL together with the administration of HMG-CoA reductase inhibitors, particularly in rodents such as hamster, in which the HMG-CoA reductase inhibitor is preferably atorvastatin.

In preferred embodiments, the antibody or antigen binding fragment thereof is characterized by one or more of the following:

(i) capable of reducing total serum cholesterol at least about 25 to about 35% and maintaining the reduction for at least a period of 24 days with respect to a predose level, preferably the reduction in total serum cholesterol is at less about 30-40%;

(ii) capable of reducing serum LDL cholesterol by at least approximately 65-80% and maintaining the reduction for at least a 24-day period with respect to a predose level;

(iii) capable of reducing serum triglyceride at least about 25-40% with respect to the predose level;

(iv) achieves one or more of (i) - (iii) without reducing serum HDL cholesterol or reducing serum HDL cholesterol no more than 5% with respect to the predose level;

(v) achieves one or more of (i) - (iii) with little or no measurable effect on liver function, as determined by measurements of ALT and AST.

In preferred embodiments, the antibody or antigen binding fragment thereof is characterized by one or more of the following:

(i) capable of reducing serum LDL cholesterol by at least approximately 40-70% and maintaining the reduction for at least a period of 60 or 90 days with respect to a predose level;

(ii) capable of reducing serum triglyceride at least about 25-40% with respect to the predose level;

(iii) does not reduce serum HDL cholesterol or reduce serum HDL cholesterol no more than 5% with respect to the predose level.

In one embodiment, the antibody or antigen binding fragment thereof is characterized by binding an epitope comprising amino acid residue 238 of PCSK9h (SEQ ID NO: 755). In a more specific embodiment, the antibody or antigen binding fragment binds an epitope comprising one or more of the amino acid residues at positions 238, 153, 159 and 343 of PCSK9h (SEQ ID NO: 755). In a more specific embodiment, the antibody or fragment thereof is characterized by binding an epitope that does not comprise an amino acid residue at positions 192, 194, 197 and / or 237 of SEQ ID NO: 755.

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In one embodiment, the antibody or antigen-binding fragment thereof is characterized by binding an epitope comprising amino acid residue 366 of PCSK9h (SEQ ID NO: 755). In a more specific embodiment, the antibody or antigen binding fragment binds an epitope comprising one or more of the amino acid residues at positions 147, 366 and 380 of PCSK9h (SEQ ID NO: 755). In a more specific embodiment, the antibody or antigen-binding fragment of an antibody is characterized by binding an epitope that does not comprise an amino acid residue at position 215 or 238 of SEQ iD NO: 755.

In one embodiment, the antibody or antigen-binding fragment thereof comprises a heavy chain variable region (HCVR) selected from the group consisting of SEQ ID NO: 2, 18, 22, 26, 42, 46, 50, 66, 70, 74, 90, 94, 98, 114, 118, 122, 138, 142, 146, 162, 166, 170, 186, 190, 194, 210, 214, 218, 234, 238, 242, 258, 262, 266, 282, 286, 290, 306, 310, 314, 330, 334, 338, 354, 358, 362, 378, 382, 386, 402, 406, 410, 426, 430, 434, 450, 454,

458, 474, 478, 482, 498, 502, 506, 522, 526, 530, 546, 550, 554, 570, 574, 578, 594, 598, 602, 618, 622, 626, 642,

646, 650, 666, 670, 674, 690, 694, 698, 714, 718, 722, 738 and 742, or a substantially similar sequence thereof having at least 90%, at least 95%, at least 98% or at least 99% sequence identity. In one embodiment, HCVR comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 50, 66, 70, 74, 90, 94, 122, 138, 142, 218, 234, 238, 242, 258, 262, 314, 330 and 334. In a more specific embodiment, HCVR comprises SEQ ID NO: 90 or 218.

In one embodiment, the antibody or antigen-binding fragment thereof further comprises a light chain variable region (LCVR) selected from the group consisting of SEQ ID NO: 10, 20, 24, 34, 44, 48, 58 , 68, 72, 82, 92, 96, 106, 116, 120, 130, 140, 144, 154, 164, 168, 178, 188, 192, 202, 212, 216, 226, 236, 240, 250, 260 , 264, 274, 284, 288, 298, 308, 312, 322, 332, 336, 346, 356, 360, 370, 380, 384, 394, 404, 408, 418, 428, 432,

442, 452, 456, 466, 476, 480, 490, 500, 504, 514, 524, 528, 538, 548, 552, 562, 572, 576, 586, 596, 600, 610, 620,

624, 634, 644, 648, 658, 668, 672, 682, 692, 696, 706, 716, 720, 730, 740 and 744, or a substantially similar sequence thereof having at least 90%, at least 95%, at least 98% or at least 99% sequence identity. In one embodiment, LCVR comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 58, 68, 72, 82, 92, 96, 130, 140, 144, 226, 236, 240, 250, 260, 264, 322, 332 and 336. In a more specific embodiment, LCVR comprises SEQ ID NO: 92 or 226.

In specific embodiments, the antibody or antigen binding fragment thereof comprises a pair of HCVR and LCVR (HCVR / LcVR) sequences selected from the group consisting of SEQ ID NO: 2/10, 18/20, 22/24 , 26/34, 42/44, 46/48, 50/58, 66/68, 70/72, 74/82, 90/92, 94/96, 98/106, 114/116, 118/120, 122 / 130, 138/140, 142/144, 146/154, 162/164, 166/168, 170/178, 186/188, 190/192, 194/202, 210/212, 214/216, 218/226 , 234/236,

238/240, 242/250, 258/260, 262/264, 266/274, 282/284, 286/288, 290/298, 306/308, 310/312, 314/322, 330/332,

334/336, 338/346, 354/356, 358/360, 362/370, 378/380, 382/384, 386/394, 402/404, 406/408, 410/418, 426/428,

430/432, 434/442, 450/452, 454/456, 458/466, 474/476, 478/480, 482/490, 498/500, 502/504, 506/514, 522/524,

526/528, 530/538, 546/548, 550/552, 554/562, 570/572, 574/576, 578/586, 594/596, 598/600, 602/610, 618/620,

622/624, 626/634, 642/644, 646/648, 650/658, 666/668, 670/672, 674/682, 690/692, 694/696, 698/706, 714/716,

718/720, 722/730, 738/740 and 742/744. In one embodiment, the sequence pair of HCVR and LCVR comprises one of SEQ ID NO: 50/58, 66/68, 70/72, 74/82, 90/92, 94/96, 122/130, 138/140 , 142/144, 218/226, 234/236, 238/240, 242/250, 258/260, 262/264, 314/322, 330/332 and 334/336. In preferred embodiments, the antibody or antigen binding fragment thereof comprises an amino acid sequence of HCVR as shown in SEQ ID NO: 90 and an amino acid sequence of LCVR as shown in SEQ ID NO: 92. In another embodiment preferred, the antibody or antigen-binding fragment thereof comprises an amino acid sequence of HCVR as shown in SEQ ID NO: 218 and an amino acid sequence of LCVR as shown in SEQ ID NO: 226.

In preferred embodiments, the antibody or antigen binding fragment thereof comprises a heavy chain CDR3 domain (HCDR3) selected from the group consisting of SEQ ID NO: 8, 32, 56, 80, 104, 128, 152, 176, 200, 224, 248, 272, 296, 320, 344, 368, 392, 416, 440, 464, 488, 512, 536, 560, 584, 608, 632, 656, 680, 704 and 728, or one substantially similar sequence thereof having at least 90%, at least 95%, at least 98% or at least 99% sequence identity; and a heavy chain CDR3 domain (LCDR3) selected from the group consisting of SEQ ID NO: 16, 40, 64, 88, 112, 136, 160, 184, 208, 232, 256, 280, 304, 328, 352 , 376, 400, 424, 448, 472, 496, 520, 544, 568, 592, 616, 640, 664, 688, 712 and 736, or substantially similar sequences thereof having at least 90%, at least 95%, at least 98% or at least 99% sequence identity. In one embodiment, the HCDR3 / LCDR3 sequence pair is selected from the group consisting of SEQ ID NO: 56/64, 80/88, 128/136, 224/232, 248/256 and 320/328. In more preferred embodiments, the antibody or antigen binding fragment thereof comprises an HCDR3 domain as shown in SEQ ID NO: 80 and an LCDR3 domain as shown in SEQ ID NO: 88. In another preferred embodiment, the antibody or the antigen binding fragment thereof comprises an HCDR3 domain as shown in SEQ ID NO: 224 and an LCDR3 domain as shown in SEQ ID NO: 232.

In a further embodiment, the antibody or antigen-binding fragment thereof further comprises a heavy chain CDR1 (HCDR1) domain selected from the group consisting of SEQ ID NO: 4, 28, 52, 76, 100, 124, 148, 172, 196, 220, 244, 268, 292, 316, 340, 364, 388, 412, 436, 460, 484, 508, 532, 556, 580, 604, 628, 652, 676, 700 and 724, or a substantially similar sequence thereof having at least 90%, at least 95%, at least 98% or at least 99% sequence identity; a heavy chain CDR2 domain

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(HCDR2) selected from the group consisting of SEQ ID NO: 6, 30, 54, 78, 102, 126, 150, 174, 198, 222, 246, 270, 294, 318, 342, 366, 390, 414, 438 , 462, 486, 510, 534, 558, 582, 606, 630, 654, 678, 702 and 726, or a substantially similar sequence thereof that has at least 90%, at least 95%, at least 98 % or at least 99% sequence identity; a light chain CDR1 domain (LCDR1) selected from the group consisting of SEQ ID NO: 12, 36, 60, 84, 108, 132, 156, 180, 204, 228, 252, 276, 300, 324, 348, 372, 396, 420, 444, 468, 492, 516, 540, 564, 588, 612, 636, 660, 684, 708 and 732, or a substantially similar sequence thereof having at least 90%, at least 95%, at least 98% or at least 99% sequence identity; and a light chain CDR2 domain (LCDR2) selected from the group consisting of SEQ ID NO: 14, 38, 62, 86, 110, 134, 158, 182, 206, 230, 254, 278, 302, 326, 350 , 374, 398, 422, 446, 470, 494, 518, 542, 566, 590, 614, 638, 662, 686, 710 and 734, or a substantially similar sequence thereof that has at least 90%, at least 95%, at least 98% or at least 99% sequence identity. In one embodiment, the CDR sequences of the heavy and light chain comprise a sequence selected from the group consisting of SEQ ID NO: 52, 54, 56, 60, 62, 64; 76, 78, 80, 84, 86, 88; 124, 126, 128, 132, 134, 136; 220, 222, 224, 228, 230, 232; 244, 246, 248, 252, 254, 256; and 316, 318, 320, 324, 326, 328. In more specific embodiments, the CDR sequences comprise SEQ ID NO: 76, 78, 80, 84, 86, 88; or 220, 222, 224, 228, 230, 232. In preferred embodiments, the antibody or antigen binding fragment thereof comprises heavy and light chain CDR amino acid sequences as shown in SEQ ID NO: 76, 78 , 80, 84, 86 and 88. In another preferred embodiment, the antibody or antigen binding fragment thereof comprises heavy and light chain CDR amino acid sequences as shown in SEQ ID NO: 220, 222, 224, 228, 230 and 232.

In a related embodiment, the antibody or antigen binding fragment thereof comprises heavy and light chain CDR domains contained within pairs of heavy and light chain sequences selected from the group consisting of SEQ ID NO: 2/10, 18/20, 22/24, 26/34, 42/44, 46/48, 50/58, 66/68, 70/72, 74/82, 90/92, 94/96, 98/106, 114 / 116, 118/120, 122/130, 138/140, 142/144, 146/154, 162/164, 166/168, 170/178, 186/188, 190/192, 194/202, 210/212, 214/216, 218/226, 234/236, 238/240, 242/250, 258/260, 262/264, 266/274, 282/284,

286/288, 290/298, 306/308, 310/312, 314/322, 330/332, 334/336, 338/346, 354/356, 358/360, 362/370, 378/380,

382/384, 386/394, 402/404, 406/408, 410/418, 426/428, 430/432, 434/442, 450/452, 454/456, 458/466, 474/476,

478/480, 482/490, 498/500, 502/504, 506/514, 522/524, 526/528, 530/538, 546/548, 550/552, 554/562, 570/572,

574/576, 578/586, 594/596, 598/600, 602/610, 618/620, 622/624, 626/634, 642/644, 646/648, 650/658, 666/668,

670/672, 674/682, 690/692, 694/696, 698/706, 714/716, 718/720, 722/730, 738/740 and 742/744. In one embodiment, the CDR sequences are contained within HCVR and LCVR selected from the amino acid sequence pairs of SEQ ID NO: 50/58, 66/68, 70/72, 74/82, 90/92, 94 / 96, 122/130, 138/140, 142/144, 218/226, 234/236, 238/240, 242/250, 258/260, 262/264, 314/322, 330/332 and 334/336. In more specific embodiments, the CDR sequences are comprised within HCVR / LCVR sequences selected from SEQ ID NO: 90/92 or 218/226. In preferred embodiments, the antibody or antigen binding fragment thereof comprises the heavy and light chain CDRs of an amino acid sequence pair of HCVR / LCVR as shown in SEQ ID NO: 90/92. In another preferred embodiment, the antibody or antigen binding fragment thereof comprises the heavy and light chain CDRs of a pair of HCVR / LCVR amino acid sequences as shown in SEQ ID NO: 218/226.

In a specific embodiment, the antibody or antigen-binding fragment thereof comprises the heavy chain variable region (HCVR) of SEQ ID NO: 90 or a substantially similar sequence thereof having at least 90%, at minus 95%, at least 98% or at least 99% sequence identity.

In a specific embodiment, the antibody or antigen-binding fragment thereof further comprises the light chain variable region (LCVR) of SEQ ID NO: 92 or a substantially similar sequence thereof having at least 90%, at least 95%, at least 98% or at least 99% sequence identity.

In specific embodiments, the antibody or antigen binding fragment thereof comprises the amino acid sequence of HCVR as shown in SEQ ID NO: 90 and an amino acid sequence of LCVR as shown in SEQ ID NO: 92.

In specific embodiments, the antibody or antigen binding fragment thereof comprises a heavy chain CDR3 domain (HCDR3) of SEQ ID NO: 80 or a substantially similar sequence thereof having at least 90%, at least 95%, at least 98% or at least 99% sequence identity; and / or a heavy chain CDR3 domain (LCDR3) of SEQ ID NO: 88, or substantially similar sequences thereof, having at least 90%, at least 95%, at least 98% or at least 99% sequence identity. In one embodiment, the sequence pair of HCDR3 / LCDR3 is SEQ ID NO: 80/88. In more preferred embodiments, the antibody or antigen binding fragment thereof comprises an HCDR3 domain as shown in SEQ ID No: 80 and an LCDR3 domain as shown in SEQ ID No: 88.

In a specific additional embodiment, the antibody or antigen binding fragment thereof further comprises the heavy chain CDR1 domain (HCDR1) of SEQ ID NO: 76, or a substantially similar sequence thereof having at least 90% , at least 95%, at least 98% or at least 99% sequence identity; and / or the CDR2 domain of the heavy chain (HCDR2) of SEQ ID NO: 78 or a substantially similar sequence thereof having at least 90%, at least 95%, at least 98% or at least 99 % sequence identity; and / or a light chain CDR1 domain (LCDR1) of SEQ ID NO: 84 or a

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substantially similar sequence thereof having at least 90%, at least 95%, at least 98% or at least 99% sequence identity; and / or a light chain CDR2 domain (LCDR2) of SEQ ID NO: 86, or a substantially similar sequence thereof having at least 90%, at least 95%, at least 98% or at least 99% sequence identity. In one embodiment, the CDR sequences of the heavy and light chain comprise a sequence selected from the group consisting of SEQ ID NO: 76, 78, 80, 84, 86, 88. In preferred embodiments, the antibody or fragment binding to the antigen thereof comprises CDR amino acid sequences of the heavy and light chain as shown in SEQ ID NO: 76, 78, 80, 84, 86 and 88.

In another specific embodiment, the antibody or antigen binding fragment thereof comprises CDR domains of the heavy and light chain contained within the pair of heavy and light chain sequences of SEQ ID NO: 90/92.

A particularly preferred embodiment relates to an antibody comprising HCVR / LCVR sequences SEQ ID NO: 90/92 and / or CDR sequences SEQ ID NO: 76, 78, 80 and / or CDR sequences SEQ ID NO: 84, 86, 88. Another particularly preferred embodiment relates to an antibody comprising the HCVR / LCVR sequences SEQ ID NO: 90/92 and the CDR sequences SEQ ID NO: 76, 78, 80 and the CDR sequences SEQ ID NO : 84, 86, 88 ("316P").

In one embodiment, the antibody or antigen binding fragment thereof comprises an HCVR encoded by a nucleic acid sequence selected from the group consisting of SEQ ID NO: 1, 17, 21, 25, 41,45, 49, 65, 69, 73, 89, 93, 97, 113, 117, 121, 137, 141, 145, 161, 165, 169, 185, 189, 193, 209, 213, 217, 233, 237, 241, 257, 261, 265, 281, 285, 289, 305, 309, 313, 329, 333, 337, 353, 357, 361, 377, 381, 385, 401, 405, 409, 425, 429, 433,

449, 453, 457, 473, 477, 481, 497, 501, 505, 521, 525, 529, 545, 549, 553, 569, 573, 577, 593, 597, 601, 617, 621,

625, 641, 645, 649, 665, 669, 673, 689, 693, 697, 713, 717, 721, 737 and 741, or a substantially identical sequence that has at least 90%, at least 95%, at less than 98%, or at least 99% sequence identity thereof. In one embodiment, HCVR is encoded by a nucleic acid sequence selected from the group consisting of SEQ ID NO: 49, 65, 69, 73, 89, 93, 121, 137, 141, 217, 233, 237, 241, 257 , 261, 313, 329 and 333. In more specific embodiments, HCVR is encoded by a nucleic acid sequence selected from the group consisting of SEQ Id NO: 89 and 217.

In one embodiment, the antibody or fragment thereof further comprises an LCVR encoded by a nucleic acid sequence selected from the group consisting of SEQ ID NO: 9, 19, 23, 33, 43, 47, 57, 67, 71, 81 , 91, 95, 105, 115, 119, 129, 139, 143, 153, 163, 167, 177, 187, 191, 201, 211, 215, 225, 235, 239, 249, 259, 263, 273, 283 , 287, 297, 307, 311, 321, 331, 335, 345, 355, 359, 369, 379, 383, 393, 403, 407, 417, 427, 431, 441, 451,

455, 465, 475, 479, 489, 499, 503, 513, 523, 527, 537, 547, 551, 561, 571, 575, 585, 595, 599, 609, 619, 623, 633,

643, 647, 657, 667, 671, 681, 691, 695, 705, 715, 719, 729, 739 and 743, or a substantially identical sequence that has at least 90%, at least 95%, at least 98%, or at least 99% sequence identity thereof. In one embodiment, the LCVR is encoded by a nucleic acid sequence selected from the group consisting of SEQ ID NO: 57, 67, 71, 81, 91, 95, 129, 139, 143, 225, 235, 239, 249, 259, 263, 321, 331 and 335. In more specific embodiments, the LCVR is encoded by a nucleic acid sequence selected from the group consisting of SEQ ID NO: 91 and 225.

In one embodiment, the antibody or antigen binding fragment thereof comprises an HCDR3 domain encoded by a nucleotide sequence selected from the group consisting of SEQ ID NO: 7, 31, 55, 79, 103, 127, 151, 175, 199, 223, 247, 271, 295, 319, 343, 367, 391, 415, 439, 463, 487, 511, 535, 559, 583, 607, 631, 655, 679, 703 and 727, or a sequence substantially identical that has at least 90%, at least 95%, at least

98%, or at least 99% sequence identity thereof; and an LCDR3 domain encoded by a nucleotide sequence selected from the group consisting of SEQ ID NO: 15, 39, 63, 87, 111, 135, 159, 183, 207, 231, 255, 279, 303, 327, 351, 375, 399, 423, 447, 471, 495, 519, 543, 567, 591, 615, 639, 663, 687, 711 and 735, or a substantially identical sequence that has at least 90%, at least 95% , at least 98%, or at least

99% sequence identity thereof. In one embodiment, HCDR3 and LCDR3 comprise a pair of sequences encoded by the nucleic acid sequence of SEQ ID NO: 55/63, 79/87, 127/135, 223/231, 247/255 and 319/327, respectively. In more specific embodiments, HCDR3 and LCDR3 comprise a pair of sequences encoded by the nucleic acid sequence of sEq ID NO: 79/87 and 223/231.

In a further embodiment, the antibody or antigen binding fragment thereof further comprises: an HCDR1 domain encoded by a nucleotide sequence selected from the group consisting of SEQ ID NO: 3, 27, 51, 75, 99, 123, 147 , 171, 195, 219, 243, 267, 291, 315, 339, 363, 387, 411,435, 459, 483, 507, 531, 555, 579, 603, 627, 651, 675, 699 and 723, or a sequence substantially identical that has at least 90%, at least 95%, at least 98%, or at least 99% sequence identity thereof; an HCDR2 domain encoded by a nucleotide sequence selected from the group consisting of SEQ ID NO: 5, 29, 53, 77, 101, 125, 149, 173, 197, 221, 245, 269, 293, 317, 341, 365 , 389, 413, 437, 461,485, 509, 533, 557, 581,605, 629, 653, 677, 701 and 725, or a substantially identical sequence that has at least 90%, at least 95%, at least 98 %, or at least 99% sequence identity thereof; an LCDR1 domain encoded by a nucleotide sequence selected from the group consisting of SEQ ID NO: 11, 35, 59, 83, 107, 131, 155, 179, 203, 227, 251, 275, 299, 323, 347, 371 , 395, 419, 443, 467, 491, 515, 539, 563, 587, 611, 635, 659, 683, 707 and 731, or a substantially identical sequence that has at least 90%, at least 95%, at least 98%, or at least 99% of

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sequence identity thereof; and an LCDR2 domain encoded by a nucleotide sequence selected from the group consisting of SEQ ID NO: 13, 37, 61, 85, 109, 133, 157, 181, 205, 229, 253, 277, 301, 325, 349, 373, 397, 421, 445, 469, 493, 517, 541, 565, 589, 613, 637, 661, 685, 709 and 733, or a substantially identical sequence that has at least 90%, at least 95% , at least 98%, or at least 99% sequence identity thereof. In one embodiment, the heavy and light chain CDR sequences are encoded by the nucleic acid sequences of SEQ ID NO: 51, 53, 55, 59, 61, 63; 75, 77, 79, 83, 85, 87; 123, 125, 127, 131, 133, 135; 219, 221, 223, 227, 229, 231; 243, 245, 247, 251, 253, 255; and 315, 317, 319, 323, 325, 327. In more specific embodiments, the heavy and light chain CDR sequences are encoded by the nucleic acid sequences of SEQ ID NO: 75, 77, 79, 83, 85 , 87; and 219, 221, 223, 227, 229, 231.

In a further embodiment, the antibody or antigen binding fragment thereof comprises an HCDR3 and an LCDR3, wherein HCDR3 comprises an amino acid sequence of formula X1-X2-X3-X4-X5-X6-X7-X8-X9 - X10 - X11 - X12 - X13 - X14 - X15 - X16 - X17 - X18 - X19 - X20 (SEQ ID NO: 747), where X1 is Ala, X2 is Arg or Lys, X3 is Asp, X4 is Ser or Ile, X5 is Asn or Val, X6 is Leu or Trp, X7 is Gly or Met, X8 is Asn or Val, X9 is Phe or Tyr, X10 is Asp, X is Leu or Met, X is Asp or is absent, X is Tyr or is absent, X is Tyr or is absent, X is Tyr

1 1 17 J 1ft J 1Q J

or is absent, X is Tyr or is absent, X is Gly or is absent, X is Met or is absent, X is Asp or is

20 1 2 3 4

absent and X is Val or absent; and LCDR3 comprises an amino acid sequence of formula X-X-X-X-X5-X6-X7-X8-X9 (SEQ ID NO: 750), wherein X1 is Gln or Met, X2 is Gln, X3 is Tyr or Thr, X4 is Tyr or Leu, X5 is Thr or Gln, X6 is Thr, X7 is Pro, X8 is Tyr or Leu and X9 is Thr.

In a further embodiment, the antibody or antigen binding fragment thereof further comprises an HCDR1 sequence of formula X1-X2-X3-X ^ -X5-X6-X7-X8 (SEQ ID NO: 745), in which X1 is Gly, X2 is Phe, X3 is Thr, X4 is Phe, X5 is Ser or Asn, X6 is Ser or Asn, X7 is Tyr or His and X8 is Ala or Trp; an HCDR2 sequence of formula X1 - X2 - X3 - X4 - X5 - X6 - X7 - X8 (SEQ ID NO: 746), in which X1 is Ile, X2 is Ser or Asn, X3 is

Gly or Gln, X4 is Asp or Ser, X5 is Gly, X6 is Ser or Gly, X7 is Thr or Glu and X8 is Thr or Lys; a sequence of LCDR1

of formula X1 - X2 - X3 - X4 - X5 - X6 - X7 - X8 - X9 - X10 - X11 - X12 (SEQ ID NO: 748) in which X1 is Gln, X2 is Ser, X3 is Val or Leu, X4 is Leu, X5 is His or Tyr, X6 is Arg or Ser, X7 is Ser or Asn, X8 is Asn or Gly, X9 is Asn, X10 is Arg or Asn, X is Asn or Tyr and X is Phe or is absent; an LCDR2 sequence of formula X-X-X

(SEQ ID NO: 749) in which X1 is Trp or Leu, X2 is Ala or Gly and X3 is Ser.

In a further embodiment, the antibody or antigen binding fragment thereof is a human anti-PCSK9 antibody or antigen binding fragment thereof comprising a heavy chain variable region (HCVR) encoded by the segments of the sequence of nucleotides derived from the germline sequences Vh, Dh and Jh, and a variable region of the light chain (LCVR) encoded by segments of the nucleotide sequence derived from germline sequences of Vk and Jk, in which the Germline sequences are (a) Vh 3-23 gene segment, Dh 7-27 gene segment, Jh 2 gene segment, Vk 4-1 gene segment and Jk 2 gene segment; or (b) Vh 3-7 gene segment, Dh 2-8 gene segment, Jh 6 gene segment, Vk 2-28 gene segment and Jk 4 gene segment.

In preferred embodiments, the antibody or antigen binding fragment thereof binds to the same epitope in PCSK9h as an antibody comprising heavy and light chain CDR amino acid sequences as shown in SEQ ID NO: 76, 78, 80 , 84, 86 and 88, or as shown in SEQ ID NO: 220, 222, 224, 228, 230 and 232.

In preferred embodiments, the antibody or antigen binding fragment thereof competes to bind PCSK9h with an antibody comprising heavy and light chain CDR amino acid sequences as shown in SEQ ID NO: 76, 78, 80, 84 , 86 and 88, or as shown in SEQ ID NO: 220, 222, 224, 228, 230 and 232.

The disclosure encompasses anti-PCSK9 antibodies that have a modified glycosylation pattern. In some applications, modification to eliminate undesirable glycosylation sites may be useful, or for example, the removal of a fucose moiety to increase the function of antibody-dependent cellular cytotoxicity (ADCC) (see Shield et al. (2002 ) JBC 277: 26733). In other applications, galactosylation modification may be made in order to modify complement dependent cytotoxicity (CDC).

Some preferred sequences related to the preferred antibodies to practice the present disclosure:

SEQ ID NO: 76: Gly Phe Thr Phe Asn Asn Tyr Ala

SEQ ID NO: 78: Ile Ser Gly Ser Gly Gly Thr Thr

SEQ ID NO: 80: Ala Lys Asp Ser Asn Trp Gly Asn Phe Asp Leu

SEQ ID NO: 84: Gln Ser Val Leu Tyr Arg Ser Asn Asn Arg Asn Phe

SEQ ID NO: 86: Trp Ala Ser

SEQ ID NO: 88: Gln Gln Tyr Tyr Thr Thr Pro Tyr Thr

5

SEQ ID NO: 90:

Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly

10

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Be Leu Arg Leu Be Cys Wing Be Ser Gly Phe Thr Phe Asn Asn

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Wing Met Asn Trp Val Arg Gln Pro Wing Gly Lys Gly Leu Asp Trp

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Ser Thr lie Ser Gly Ser Gly Gly Thr Thr Asn Tyr Ala Asp Ser

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Lys Gly Arg Phe lie lie Ser Arg Asp Ser Ser Lys His Thr Leu

65

70

75

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr A.la Val Tyr Tyr

85

90

95

Wing Lys Asp Ser Asn Trp Gly Asn Phe Asp Leu Trp Gly Arg Gly

100

Leu Val Thr Val Ser Ser 115

105

110

SEQ ID NO: 92:

image 1

SEQ ID NO: 755 (PCSK9h):

Gly

Tyr

Val

Val

Tyr

80

Cys

Thr

Gly

Arg

Gln

Val

Thr

80

Gln

lie

Claims (16)

5 10 fifteen twenty 25 30 35 40 Four. Five fifty 1. Pharmaceutical composition comprising an antibody or an antigen-binding fragment thereof that specifically binds to proprotein convertase subtilisin / human type 9 kexin (PCSK9h) together with a pharmaceutically acceptable excipient or vehicle for use in the treatment of a disease or condition, in which the expression or activity of PCSK9 causes an impact, in a human patient, wherein the antibody or antigen binding fragment thereof comprises the three heavy chain CDRs set forth in SEQ ID NO: 76, 78, and 80 and the three light chain CDRs set forth in SEQ ID NO: 84, 86 , and 88, wherein the pharmaceutical composition is comprised in a unit dosage form, in which the unit dosage form comprises 75 mg, 150 mg or 300 mg of the antibody or antigen binding fragment thereof, in which the pharmaceutical composition is administered every two to four weeks, and in which the disease or condition is selected from the group consisting of high levels of total cholesterol, high levels of non-high-density lipoprotein (HDL) cholesterol, high levels of low-density lipoprotein (C-LDL), high levels of apolipoprotein B100 (ApoB100), hyperlipidemia, dyslipidemia, atherosclerosis, cardiovascular diseases, such as coronary heart disease (ECC) and hypercholesterolemia, such as primary hypercholesterolemia, familial hypercholesterolemia, unfamiliar hypercholesterolemia, heterozygous familial hypercholesterolemia (heFH), noncholesterolemia controlled by statins, and hypercholesterolemia that is resistant to statins. 2. Pharmaceutical composition for use according to claim 1, wherein the antibody or antigen-binding fragment thereof comprises the amino acid sequence of the heavy chain variable region (HCVR) and the amino acid sequence of the variable region of the light chain (LCVR) exposed sEq ID NO: 90 and 92, respectively. 3. Pharmaceutical composition for use according to claim 1 or 2, wherein the antibody or antigen binding fragment thereof is for administration to a subject that is in at least one of the following groups of subjects: (i) subjects who have a serum LDL (C-LDl) cholesterol level of at least 100 mg / dl, (ii) subjects who have a serum HDL-C level below 40 mg / dl; (iii) subjects who have a serum cholesterol level of at least 200 mg / dl; (iv) subjects who have a serum triacylglycerols level of at least 150 mg / dl, in which said triacylglycerols level is determined after fasting for at least 8 hours. 4. Pharmaceutical composition for use according to one of claims 1 to 3, comprising an effective dose of the antibody or antigen binding fragment thereof, wherein the dose is sufficient for sustained reduction of low density lipoprotein levels. (C-LDL) for a period of at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23 or at At least 28 days after administration. 5. Pharmaceutical composition for use according to one of claims 1 to 4, wherein the antibody or antigen binding fragment thereof has one or more of the following characteristics: (a) reduction of LDL-C from at least -60% to at least -75% with respect to a pre-dose level with a sustained reduction for at least a period of 14 days after administering a dose of 150 mg to a subject E2W; (b) increase in serum HDL cholesterol levels of at least 2%, at least 2.5%, at least 3%, at least 3.5%, at least 4%, at least 4.5%, at least 5% or at least 5.5% with respect at a predose level after administering a dose of 150 mg E2W to a subject; (c) reduction of one or more of: total cholesterol levels, ApoB levels, C-non HDL levels, Apo-B / ApoA-1 ratio, after administration to a subject. 6. Pharmaceutical composition for use according to one of claims 1 to 5 comprising the antibody or antigen binding fragment thereof as a liquid formulation. 7. Pharmaceutical composition for use according to claim 6, wherein the liquid formulation is a solution for injection comprising 75 mg of the antibody or antigen binding fragment thereof per 1 ml of volume. 5 10 fifteen twenty 25 30 35 8. Pharmaceutical composition for use according to claim 6, wherein the liquid formulation is a solution for injection comprising 150 mg of the antibody or antigen-binding fragment thereof per 1 ml of volume. 9. Pharmaceutical composition for use according to claim 1, wherein the unit dosage form comprises 75 mg of the antibody or antigen binding fragment thereof. 10. Pharmaceutical composition for use according to claim 1, wherein the unit dosage form comprises 150 mg of the antibody or antigen binding fragment thereof. 11. Pharmaceutical composition for use according to claim 1, wherein the unit dosage form comprises 300 mg of the antibody or antigen binding fragment thereof. 12. Pharmaceutical composition for use according to one of claims 1 to 11, wherein the unit dosage form comprises the antibody or antigen binding fragment thereof as a liquid formulation in a hermetically sealed container selected from the group consisting of a vial, an envelope, a pre-filled syringe, a pre-filled auto-injector, a cartridge for a reusable syringe and an applicator. 13. Pharmaceutical composition for use according to one of claims 1 to 12, wherein the pharmaceutical composition is administered by subcutaneous injection. 14. Pharmaceutical composition for use according to one of claims 1 to 13, wherein the antibody or antigen binding fragment thereof binds an epitope comprising one or more of the amino acid residues at positions 238, 153, 159 and 343 of PCSK9h (SEQ ID NO: 755). 15. Pharmaceutical composition comprising 75 mg of an antibody or antigen-binding fragment thereof that specifically binds to proprotein convertase subtilisin / human type 9 kexin (PCSK9h) together with a pharmaceutically acceptable carrier or excipient for use according to claim one, wherein the antibody or antigen-binding fragment thereof comprises the amino acid sequence of the heavy chain variable region (HCVR) and the amino acid sequence of the light chain variable region (LCVR) set forth in SEQ ID NO : 90 and 92, respectively, in which the pharmaceutical composition is a solution for injection of 1 ml, in which the pharmaceutical composition is administered every two to four weeks. 16. Pharmaceutical composition comprising 150 mg of an antibody or an antigen-binding fragment thereof that specifically binds to proprotein convertase subtilisin / human type 9 kexin (PCSK9h) together with a pharmaceutically acceptable excipient or vehicle for use according to claim one, wherein the antibody or antigen-binding fragment thereof comprises the amino acid sequence of the heavy chain variable region (HCVR) and the amino acid sequence of the light chain variable region (LCVR) set forth in SEQ ID NO : 90 and 92, respectively, in which the pharmaceutical composition is a solution for injection of 1 ml, in which the pharmaceutical composition is administered every two to four weeks. Days % of variation with respect to the initial level image 1 I §íá image2 10 or 10 twenty Trj cíq " to 100 mg-FH-Yes __ FBG-FH-No ““ PBG-FH-Yes 100 mg-combined PBO-combined image3 i— | —i — i — i — i— | —i — i — i — i— | —i — i — i — i— | —i — i — i — i— | —i — i — i— i— | —i — i — i — i — i- 30 40 50 60 70 80 90 Days Days % of variation with respect to the initial level image4 and (Open preinclusion: atorvastatin) image5 Selection Visit Selection period 7 weeks Treatment period (8 weeks) FU period (8 weeks) -------------------------------------------------- ----------------------------- ^ image6 >

 to S 2

 S 4 A S 6 S 8 S 12 S 16

 D 15

 D 29 D 43 D 57 D 85 D 113

* NCEP-ATPIII TLC or equivalent diet oo 1 week selection period Selection Visit image7 s -1 D -7 S 0 D 1 Diet N = 30 N = 30

 N = 30

 316P + atorvastatin 10 mg

 (maintenance dose)

image8 00 00 image9 Placebo 50mgE2W 200mgE4W 300mgE4W 100mgE2W 150mgE2W